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H I G H -Y I E L D P R I N C I P L E S I N

Biochemistry
“Biochemistry is the study of carbon compounds that crawl.” ! High-Yield Clinical
––Mike Adams Vignettes

! Molecular

! Cellular

! Metabolism
This high-yield material includes molecular biology, genetics, cell bi-
ology, and principles of metabolism (especially vitamins, cofactors, ! Laboratory Techniques
minerals, and single-enzyme-deficiency diseases). When studying ! Genetics
metabolic pathways, emphasize important regulatory steps and en-
zyme deficiencies that result in disease. For example, understanding ! Nutrition
the defect in Lesch-Nyhan syndrome and its clinical consequences is
higher yield than memorizing every intermediate in the purine sal-
vage pathway. Do not spend time on hard-core organic chemistry,
mechanisms, and physical chemistry. Detailed chemical structures
are infrequently tested. Familiarity with the latest biochemical tech-
niques that have medical relevance––such as enzyme-linked im-
munosorbent assay (ELISA), immunoelectrophoresis, Southern blot-
ting, and PCR––is useful. Beware if you placed out of your medical
school’s biochemistry class, for the emphasis of the test differs from
that of many undergraduate courses. Review the related biochemistry
when studying pharmacology or genetic diseases as a way to reinforce
and integrate the material.

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B I O C H E M I ST RY—H I G H-Y I E LD C LI N I C AL V I G N E T T E S
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! Full-term neonate of uneventful What is the diagnosis? PKU.


delivery becomes mentally
retarded and hyperactive and
has a musty odor.
! Stressed executive comes What is the mechanism? NADH increase prevents
home from work, consumes 7 gluconeogenesis by shunting
or 8 martinis in rapid succession pyruvate and oxaloacetate to
before dinner, and becomes lactate and malate.
hypoglycemic.
! 2-year-old girl has an ↑ in What is the diagnosis? Kwashiorkor.
abdominal girth, failure to thrive,
and skin and hair depigmentation.
! Alcoholic develops a rash, What is the vitamin Vitamin B3 (pellagra).
BIOC H E M ISTRY

diarrhea, and altered mental deficiency?


status.
! 51-year-old man has black spots What is the diagnosis? Alkaptonuria.
in his sclera and has noted that
his urine turns black upon
standing.
! 25-year-old male complains What is the disease, Familial hypercholesterolemia;
of severe chest pain and has and where is the defect? LDL receptor.
xanthomas of his Achilles
tendons.

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Chromatin Condensed by (−) charged DNA looped twice Think of beads on a string.
structure around (+) charged H2A, H2B, H3, and H4
histone octamers (nucleosome bead). H1 ties
nucleosomes together in a string (30-nm fiber).
In mitosis, DNA condenses to form mitotic
chromosomes.
Heterochromatin Condensed, transcriptionally inactive.
Euchromatin Less condensed, transcriptionally active. Eu = true, “truly transcribed.”

DNA
Histone H1

Nucleosome core

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histones H2A, H2B, H3, H4

Nucleotides Purines (A, G) have 2 rings. Pyrimidines (C, T, U) PURe As Gold: PURines.
have 1 ring. Guanine has a ketone. Thymine has CUT the PY (pie):
a methyl. Deamination of cytosine makes uracil. PYrimidines.
Uracil found in RNA; thymine in DNA. THYmine has a meTHYl.
G-C bond (3 H-bonds) stronger than A-T bond
(2 H-bonds). ↑ G-C content → ↑ melting
temperature.

Purine (A, G) Pyrimidine (C, T, U)

Carbamoyl
CO2
phosphate
Glycine
Aspartate
C N C
N C N C
C N10–Formyl-
C C tetrahydrofolate C C
N N N
N 10–Formyl-
Glutamine
tetrahydrofolate Aspartate
Nucleotides are linked by 3′-5′ phosphodiesterase bond.

Transition vs. Transition––substituting purine for purine or TransItion = Identical type.


transversion pyrimidine for pyrimidine.
Transversion––substituting purine for pyrimidine TransVersion = conVersion
or vice versa. between types.

Genetic code Unambiguous––each codon specifies only 1 amino acid.


features Degenerate––more than 1 codon may code for same amino acid.
Commaless, nonoverlapping (except some viruses).
Universal (exceptions include mitochondria, archaeobacteria, Mycoplasma, and some
yeasts).

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Mutations in DNA Silent––same aa, often base change in 3rd position Severity of damage: nonsense
of codon (tRNA wobble). > missense > silent.
Missense––changed aa (conservative––new aa is
similar in chemical structure).
Nonsense––change resulting in early stop codon. Stop the nonsense!
Frame shift––change resulting in misreading of all
nucleotides downstream, usually resulting in a
truncated protein.

Prokaryotic DNA Single origin of replication––continuous DNA DNA polymerase III has
replication and synthesis on leading strand and discontinuous 5′ → 3′ synthesis and
DNA polymerases (Okazaki fragments) on lagging strand. proofreads with 3′ → 5′
DNA topoisomerases create a nick in the helix to exonuclease.
relieve supercoils. DNA polymerase I excises
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Primase makes an RNA primer on which DNA RNA primer with 5′ → 3′


polymerase III can initiate replication. exonuclease.
DNA polymerase III elongates the chain by adding
deoxynucleotides to the 3′ end until it reaches
primer of preceding fragment. 3′ → 5′
exonuclease activity “proofreads” each added
nucleotide.
DNA polymerase I degrades RNA primer.
DNA ligase seals.

DNA 3'
polymerase III
5'
Single-strand
binding protein Leading strand

RNA
primer
DNA Okazaki
polymerase III fragment DNA
ligase
5'
Lagging
strand

3' Primase 3'

5'

Eukaryotic DNA Eukaryotic genome has multiple origins of replication. Replication begins at a
polymerases consensus sequence of AT base pairs.
Eukaryotes have separate polymerases (α, β, γ, δ, ε) for synthesizing RNA primers,
leading-strand DNA, lagging-strand DNA, mitochondrial DNA, and DNA repair.

DNA repair: single Single-strand, excision repair–specific glycosylase recognizes and removes damaged
strand base. Endonuclease makes a break several bases to the 5′ side. Exonuclease removes
short stretch of nucleotides. DNA polymerase fills gap. DNA ligase seals.

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DNA repair defects Xeroderma pigmentosum (skin sensitivity to UV light), ataxia-telangiectasia (x-rays),

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Bloom’s syndrome (radiation), and Fanconi’s anemia (cross-linking agents).

Xeroderma Defective excision repair such as uvr ABC Autosomal recessive.


pigmentosum endonuclease. Results in inability to repair
thymidine dimers, which form in DNA when Thymidine UV
exposed to UV light. dimer
3' 5'
Associated with dry skin and with melanoma and
5' 3'
other cancers.

DNA/RNA/protein DNA and RNA are both synthesized 5′ → 3′. Imagine the incoming
synthesis direction Remember that the 5′ of the incoming nucleotide nucleotide bringing a gift
bears the triphosphate (energy source for bond). (triphosphate) to the 3′ host.
The 3′ hydroxyl of the nascent chain is the target. “BYOP (phosphate) from 5
Protein synthesis also proceeds in the 5′ to 3′ to 3.”
direction. Amino acids are linked N

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to C.

Types of RNA mRNA is the largest type of RNA. Massive, Rampant, Tiny.
rRNA is the most abundant type of RNA.
tRNA is the smallest type of RNA.

RNA polymerases
Eukaryotes RNA polymerase I makes rRNA. I, II, and III are numbered as
RNA polymerase II makes mRNA. their products are used in
RNA polymerase III makes tRNA. protein synthesis.
No proofreading function, but can initiate chains. α-amanitin is found in death
RNA polymerase II opens DNA at promoter site cap mushrooms.
(A-T-rich upstream sequence––TATA and CAAT).
α-amanitin inhibits RNA polymerase II.
Prokaryotes RNA polymerase makes all 3 kinds of RNA.

Start and stop AUG (or rarely GUG) is the mRNA initiation codon. AUG inAUGurates
codons Eukaryotes––AUG codes for methionine, which protein synthesis.
may be removed before translation is completed.
Prokaryotes––the initial AUG codes for a formyl-
methionine (f-met).
Stop codons––UGA, UAA, UAG. UGA = U Go Away.
UAA = U Are Away.
UAG = U Are Gone.

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Regulation of gene expression


Promoter Site where RNA polymerase and multiple other Promoter mutation commonly
transcription factors bind to DNA upstream from results in dramatic ↓ in
gene locus. amount of gene transcribed.
Enhancer Stretch of DNA that alters gene expression by binding
transcription factors. May be located close to, far
from, or even within (in an intron) the gene whose
expression it regulates.
Operator Site where negative regulators (repressors) bind.

Introns vs. Exons contain the actual genetic information INtrons stay IN the nucleus,
exons coding for protein. whereas EXons EXit and are
Introns are intervening noncoding segments of DNA. EXpressed.
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Introns

DNA

Exons
Transcription
and splicing

mRNA

Splicing of mRNA Introns are precisely spliced out of 1° mRNA transcripts. A lariat-shaped intermediate
is formed. Small nuclear ribonucleoprotein particles (snRNP) facilitate splicing by
binding to 1° mRNA transcripts and forming spliceosomes.

RNA processing Occurs in nucleus. After transcription: Only processed RNA is


(eukaryotes) 1. Capping on 5′ end (7-methyl-G) transported out of the
Cap
2. Polyadenylation on 3′ end (≈ 200 A’s) nucleus.
Coding
5' 3. Splicing out of introns
Gpppp
Initial transcript is called heterogeneous nuclear
3'
HO-AAAA RNA (hnRNA).
Tail Capped and tailed transcript is called mRNA.

tRNA structure 75–90 nucleotides, cloverleaf form, anticodon end is opposite 3′ aminoacyl end. All
Methionine tRNAs, both eukaryotic and prokaryotic, have CCA at 3′ end along with a high
ACC
3'
percentage of chemically modified bases. The amino acid is covalently bound to the 3′
5' end of the tRNA.

Anticodon
(CAU) UAC

AUG
5' 3'
mRNA Codon

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tRNA charging Aminoacyl-tRNA synthetase (1 per aa, uses ATP) Aminoacyl-tRNA synthetase

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scrutinizes aa before and after it binds to tRNA. If and binding of charged
incorrect, bond is hydrolyzed by synthetase. The tRNA to the codon are
aa-tRNA bond has energy for formation of peptide responsible for accuracy of
bond. A mischarged tRNA reads usual codon but amino acid selection.
inserts wrong amino acid.
OH Aminoacyl tRNA AA
3'
synthetase
5'
ATP AMP + PPi

tRNA wobble Accurate base pairing is required only in the first 2 nucleotide positions of an mRNA

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codon, so codons differing in the 3rd “wobble” position may code for the same
tRNA/amino acid.

Protein synthesis Met sits in the P site––peptidyl. The incoming ATP––tRNA Activation
Ribosome
60S amino acid binds to the A site––aminoacyl, (charging).
hydrolyzing Met’s bond to its tRNA while GTP––tRNA Gripping and
E P A
5' 3'
simultaneously forming a peptidyl bond between Going places (translocation).
the 2 amino acids. The ribosome shifts 1 codon
40S
toward the 3′ end of the mRNA, shifting the
uncharged tRNA into the E position and the
dipeptidyl tRNA into the P site.

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Enzyme kinetics

The lower the Km, the higher

Velocity (V)
Vmax the affinity.
1
Km = [S] at Vmax
1 2
Vmax Km
2

[S]

1 Km
1 V slope =
Vmax
−K m 1
Vmax
1
[S]

Noncompetitive inhibitor
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Competitive inhibitor
HINT: Competitive inhibitors
1
V Uninhibited cross each other
competitively, while
noncompetitive inhibitors
1 do not.
[S]

Competitive Noncompetitive
inhibitors inhibitors
Resemble substrate Yes No
Overcome by ↑ [S] Yes No
Bind active site Yes No
Effect on Vmax Unchanged ↓
Effect on Km ↑ Unchanged

Enzyme regulation Enzyme concentration alteration (synthesis and/or destruction), covalent modification
methods (e.g., phosphorylation), proteolytic modification (zymogen), allosteric regulation (e.g.,
feedback inhibition), and transcriptional regulation (e.g., steroid hormones).

Cell cycle phases M (mitosis: prophase–metaphase– G stands for Gap or Growth; S


anaphase–telophase) for Synthesis.
G1 (growth)
S (synthesis of DNA)
Mitosis
G2 (growth)
G2
G0 (quiescent G1 phase)
Interphase
G1 and G0 are of variable duration. Mitosis is (G1, S, G2)
G1
usually shortest phase. Most cells are in G0. S
phase
Rapidly dividing cells have a shorter G1.

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Rough RER is the site of synthesis of secretory (exported) Mucus-secreting goblet cells of

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endoplasmic proteins and of N-linked oligosaccharide addition the small intestine and
reticulum (RER) to many proteins. antibody-secreting plasma
cells are rich in RER.

Nissl bodies Nissl bodies (in neurons)––rough ER; not found in axon or axon hillock.
Synthesize enzymes (e.g., ChAT) and peptide neurotransmitters.

Smooth SER is the site of steroid synthesis and detoxification Liver hepatocytes and
endoplasmic of drugs and poisons. steroid hormone–producing
reticulum (SER) cells of the adrenal cortex
are rich in SER.

Functions of Golgi 1. Distribution center of proteins and lipids from I-cell disease is caused by the
apparatus ER to the plasma membrane, lysosomes, and failure of addition of
secretory vesicles mannose-6-phosphate to

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2. Modifies N-oligosaccharides on asparagine lysosome proteins, causing
3. Adds O-oligosaccharides to serine and threonine these enzymes to be secreted
residues outside the cell instead of
4. Proteoglycan assembly from proteoglycan core being targeted to the
proteins lysosome. Characterized by
5. Sulfation of sugars in proteoglycans and of coarse facial features and
selected tyrosine on proteins restricted joint movement.
6. Addition of mannose-6-phosphate to specific
lysosomal proteins, which targets the protein
to the lysosome

RER

SER

Cell
membrane

Cell
membrane

cis
trans face
face

(Reproduced, with permission, from Junqueira L, Carneiro J. Basic Histology, 10th ed.
New York:McGraw-Hill, 2003.)

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Microtubule Cylindrical structure 24 nm in diameter and of variable Drugs that act on microtubules:
length. A helical array of polymerized dimers of α- 1. Mebendazole/thiabendazole
and β-tubulin (13 per circumference). Each dimer (antihelminthic)
has 2 GTP bound. Incorporated into flagella, cilia, 2. Taxol (anti–breast cancer)
mitotic spindles. Grows slowly, collapses quickly. 3. Griseofulvin (antifungal)
24 nm
Microtubules are also involved in slow axoplasmic 4. Vincristine/vinblastine
transport in neurons. (anti-cancer)
5. Colchicine (anti-gout)
Chédiak-Higashi syndrome
is due to a microtubule
polymerization defect
resulting in ↓ phagocytosis.

Cilia structure 9 + 2 arrangement of microtubules. Kartagener’s syndrome is due


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Dynein is an ATPase that links peripheral to a dynein arm defect,


9 doublets and causes bending of cilium by resulting in immotile cilia.
differential sliding of doublets. Dynein = retrograde.
Kinesin = anterograde.
Microtubule
doublets

Dynein ATPase

Plasma membrane Plasma membranes contain cholesterol (≈ 50%, promotes membrane stability),
composition phospholipids (≈ 50%), sphingolipids, glycolipids, and proteins. High cholesterol or
long saturated fatty acid content → ↑ melting temperature. Only noncytoplasmic side
of membrane contains glycosylated lipids or proteins (i.e., the plasma membrane is an
asymmetric, fluid bilayer).

Phosphatidylcholine Phosphatidylcholine (lecithin) is a major component of RBC membranes, of myelin, of


function bile, and of surfactant (DPPC––dipalmitoyl phosphatidylcholine). Also used in
esterification of cholesterol (LCAT is lecithin-cholesterol acyltransferase).

Sodium pump Na+-K+ATPase is located in the plasma membrane Ouabain inhibits by binding to
with ATP site on cytoplasmic side. For each ATP K+ site. Cardiac glycosides
consumed, 3 Na+ go out and 2 K+ come in. (digoxin, digitoxin) also
During cycle, pump is phosphorylated. inhibit the Na+-K+ATPase,
causing ↑ cardiac contractility.
Extracellular 3Na+ 2K+
side

P
Cytosolic ADP 2K+
side 3Na+ ATP P

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G-protein-linked 2nd messengers

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Receptor G-protein class
Lipids
H1, α1, V1, Gq HAVe 1 M&M.
Receptor Phospholipase C
M1, M3 IP3 [Ca2+]in
PIP2
DAG Protein
kinase C
β1, β2, D1,
Receptor ATP
H2, V2 G
s
+
Adenylyl cyclase
Gi –

M2, α2, D2 Receptor cAMP Protein kinase A MAD 2s.

Collagen types Collagen is the most abundant protein in the Be Cool, Read Books.

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human body. Functions to organize and
strengthen extracellular matrix.
Type I (90%)––Bone, tendon, skin, dentin, fascia, Type I: BONE.
cornea, late wound repair.
Type II––Cartilage (including hyaline), vitreous Type II: carTWOlage.
body, nucleus pulposus.
Type III (Reticulin)––skin, blood vessels, uterus,
fetal tissue, granulation tissue.
Type IV––Basement membrane or basal lamina. Type IV: Under the floor
Type X––epiphyseal plate. (basement membrane).

Collagen synthesis Inside fibroblasts:


and structure 1. Collagen α chains (preprocollagen)
Nucleus
translated on RER––usually Gly-X-Y
DNA
polypeptide (X and Y are proline,
hydroxyproline, or hydroxylysine)
mRNA
2. ER → hydroxylation of specific proline ER
Hydroxylation
and lysine residues (requires vitamin C) OH OH
3. Golgi → glycosylation of pro-α-chain Scurvy

lysine residues and formation of


procollagen (triple helix of 3 collagen Golgi
Glycosylation
(pro α chain)
OH OH
α chains) Osteogenesis
4. Procollagen molecules are exocytosed imperfecta

into extracellular space Triple helix (procollagen)

Outside fibroblasts: Cell membrane


c(1-)
5. Procollagen peptidases cleave terminal Peptide cleavage

regions of procollagen, transforming Ehlers-Danlos

procollagen into insoluble tropocollagen Collagen fibrils


with crosslinks
6. Many staggered tropocollagen molecules
are reinforced by covalent lysine-hydroxylysine
cross-linkage (by lysyl oxidase) to make collagen
fibrils

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Ehlers-Danlos Faulty collagen synthesis causing:


syndrome 1. Hyperextensible skin
2. Tendency to bleed (easy bruising)
3. Hypermobile joints
10 types. Inheritance varies. Associated with berry aneurysms.

Osteogenesis Primarily an autosomal-dominant disorder caused May be confused with child


imperfecta by a variety of gene defects, resulting in abnormal abuse.
collagen synthesis. Clinically characterized by: Type II is fatal in utero and in
1. Multiple fractures occurring with minimal the neonatal period.
trauma (brittle bone disease), which may Incidence is 1:10,000.
occur during the birth process
2. Blue sclerae due to the translucency of the
connective tissue over the choroid
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3. Hearing loss (abnormal middle ear bones)


4. Dental imperfections due to lack of dentition

Immunohistochemical stains
Stain Cell type
Vimentin Connective tissue
Desmin Muscle
Cytokeratin Epithelial cells
Glial fibrillary acid proteins (GFAP) Neuroglia
Neurofilaments Neurons

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Metabolism sites
Mitochondria Fatty acid oxidation (β-oxidation), acetyl-CoA production, Krebs cycle.
Cytoplasm Glycolysis, fatty acid synthesis, HMP shunt, protein synthesis (RER), steroid synthesis
(SER).
Both Gluconeogenesis, urea cycle, heme synthesis.

Summary of pathways

Galactose
1
Galactose-1-phosphate
Glycogen Glucose
3 4
2
5
UDP-glucose Glucose-1-phosphate Glucose-6-phosphate 6-phosphogluconolactone

1 Galactokinase (mild galactosemia) HMP shunt

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2 Galactose-1-phosphate uridyltransferase 6
(severe galactosemia) Fructose-6-phosphate Ribulose-5-phosphate
3 Hexokinase/glucokinase
7 8
4 Glucose-6-phosphatase (von Gierke’s)
5 Glucose-6-phosphate dehydrogenase (G6PD) Fructose-1,6-bisphosphate
6 Transketolase
7 Phosphofructokinase
8 Fructose-1,6-bisphosphatase Glyceraldehyde-3-P DHAP 10 9
F1P Fructose
9 Fructokinase (essential fructosuria)
10 Aldolase B (fructose intolerance) 1,3-bis-phosphoglycerate Glyceraldehyde
11 Pyruvate kinase
12 Pyruvate dehydrogenase
3-phosphoglycerate
13 HMG-CoA reductase
14 Pyruvate carboxylase Glycolysis
15 PEP carboxykinase 2-phosphoglycerate
16 Citrate synthase
17 α-ketoglutarate dehydrogenase Cholesterol
Phosphoenolpyruvate (PEP)
18 Ornithine transcarbamylase
11
Pyruvate Lactate
13 Mevalonate
Gluconeogenesis 12
15 Acetoacetyl-CoA HMG-CoA
Acetyl-CoA
Acetoacetate β-hydroxybutyrate
14 Malonyl-CoA Fatty acids
NH4 + CO2
Aspartate
16 Citrate
Citrulline Oxaloacetate
Carbamoyl
phosphate Isocitrate
18 Argininosuccinate
Malate TCA
Ornithine Urea
cycle
cycle
Fumarate α-ketoglutarate
17
Arginine Succinate Succinyl-CoA Methylmalonyl-CoA
Urea
Propionyl-CoA
H2O

Odd-chain
fatty acids

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ATP Base (adenine), ribose, 3 phosphoryls. 2 phosphoanhydride bonds, 7 kcal/mol each.


Aerobic metabolism of glucose produces 38 ATP via malate shuttle, 36 ATP via G3P shuttle.
NH2 Anaerobic glycolysis produces only 2 net ATP per glucose molecule.
N
N
ATP hydrolysis can be coupled to energetically unfavorable reactions.
O O O
N N
O P O
~ P O
~ P O-

O O- O- O-

OH OH

Activated carriers Phosphoryl (ATP).


Electrons (NADH, NADPH, FADH2).
Acyl (coenzyme A, lipoamide).
CO2 (biotin).
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1-carbon units (tetrahydrofolates).


CH3 groups (SAM).
Aldehydes (TPP).
Glucose (UDP-glucose).
Choline (CDP-choline).

S-adenosyl- ATP + methionine → SAM. SAM transfers methyl SAM the methyl donor man.
methionine units to a wide variety of acceptors (e.g., in
synthesis of phosphocreatine, a high-energy
phosphate active in muscle ATP production).
Regeneration of methionine (and thus SAM) is
dependent on vitamin B12.

Signal molecule ATP → cAMP via adenylate cyclase.


precursors GTP → cGMP via guanylate cyclase.
Glutamate → GABA via glutamate decarboxylase (requires vitamin B6).
Choline → ACh via choline acetyltransferase (ChAT).
Arachidonate → prostaglandins, thromboxanes, leukotrienes via cyclooxygenase/
lipoxygenase.
Fructose-6-P → fructose-1,6-bis-P via phosphofructokinase (PFK), the rate-limiting
enzyme of glycolysis.
1,3-BPG → 2,3-BPG via bisphosphoglycerate mutase.

Universal electron Nicotinamides (NAD+, NADP+) and flavin NADPH is a product of the
acceptors nucleotides (FAD+). HMP shunt.
NAD+ is generally used in catabolic processes to
carry reducing equivalents away as NADH.
NADPH is used in anabolic processes (steroid NADPH is used in:
and fatty acid synthesis) as a supply of reducing 1. Anabolic processes
equivalents. 2. Respiratory burst
3. P-450

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Oxygen-dependent respiratory burst

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Phagolysosome
Neutrophil
cell membrane

NADPH
O2
↓ 1
NADP+
O2•
2

Cl- H2O2 H2O2 H2O
↓ 3
HOCl• GSH 4 GSSG
Bacteria
1 NADPH oxidase (deficiency = ↓
chronic granulomatous disease) 5
NADP+ NADPH
2 Superoxide dismutase
3 Myeloperoxidase 6
G6P 6PG
4 Catalase

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5 Glutathione reductase GSH/GSSG = glutathione
6 Glucose-6-phosphate (reduced/oxidized)
dehydrogenase (G6PD) HOCl• = bleach

Hexokinase vs. Hexokinase is found throughout body. Only hexokinase is feedback


glucokinase GLucokinase is primarily found in the Liver inhibited by G6P.
(lower affinity [↑ Km] but higher capacity Glucokinase phosphorylates
[↑ Vmax]). excess glucose (e.g., after a
meal) to sequester it in the
liver as G6P.

Glycolysis D-glucose Glucose-6-phosphate Glucose-6-P ".


regulation, Hexokinase/glucokinase*
irreversible Fructose-6-P Fructose-1,6-BP ATP ", AMP ⊕, citrate ",
enzymes Phosphofructokinase-1 fructose-2,6-BP ⊕.
(rate-limiting step)

Phosphoenolpyruvate Pyruvate ATP ", alanine ",


Pyruvate kinase fructose-1,6-BP ⊕.
Pyruvate Acetyl-CoA ATP ", NADH ",
Pyruvate acetyl-CoA ".
dehydrogenase

* Glucokinase in liver; hexokinase in all other tissues.

Glycolytic enzyme Hexokinase, glucose phosphate isomerase, aldolase, RBCs metabolize glucose
deficiency triosephosphate isomerase, phosphate glycerate anaerobically (no
kinase, enolase, and pyruvate kinase deficiencies mitochondria) and thus
are associated with hemolytic anemia. depend solely on glycolysis.

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Pyruvate The complex contains 3 enzymes that require 5 The complex is similar to the
dehydrogenase cofactors (the first 4 B vitamins plus lipoic acid): α-ketoglutarate
complex 1. Pyrophosphate (B1, thiamine; TPP) dehydrogenase complex
2. FAD (B2, riboflavin) (same cofactors, similar
3. NAD (B3, niacin) substrate and action).
4. CoA (B5, pantothenate)
5. Lipoic acid
Reaction: pyruvate + NAD+ + CoA → acetyl-CoA +
CO2 + NADH.
Activated by exercise:
↑ NAD+/NADH ratio
↑ ADP
↑ Ca2+
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Pyruvate Causes backup of substrate (pyruvate and alanine), Lysine and Leucine––the only
dehydrogenase resulting in lactic acidosis. Can be seen in purely ketogenic amino
deficiency alcoholics due to B1 deficiency. acids.
Findings: neurologic defects.
Treatment: ↑ intake of ketogenic nutrients (e.g., high
fat content or ↑ lysine and leucine).

Pyruvate metabolism 6 ATP equivalents are needed


Glucose to generate glucose from
Cytosol
pyruvate.
Alanine
ALT
Pyruvate
LDH
Lactate
Alanine serves as carrier of
amino groups from muscle to
NADH + H+ NAD+
PC liver.
CO2 + ATP NAD+ PDH
Oxaloacetate can be used to
CO2
replenish TCA cycle or in
NADH + H+ gluconeogenesis.
Oxaloacetate Acetyl-CoA

Mitochondria

Cori cycle Transfers excess reducing


equivalents from RBCs and
MUSCLE LIVER
muscle to liver, allowing
Glucose Glucose
muscle to function
2ATP B 6ATP
Pyruvate L Pyruvate anaerobically (net 2 ATP).
O
Lactate O Lactate
dehydrogenase D dehydrogenase

Lactate Lactate

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TCA cycle Produces 3 NADH, 1 FADH2,

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Pyruvate
- ATP 2 CO2, 1 GTP per acetyl-
Pyruvate
dehydrogenase
- Acetyl-CoA CoA = 12 ATP/acetyl-CoA
- NADH
Acetyl-CoA (2× everything per glucose)
- ATP

Oxalo-
α-ketoglutarate dehydrogenase
Citrate Citrate
NADH
acetate synthase complex requires same
cis-aconitate cofactors as the pyruvate
Malate
Isocitrate CO2 + NADH dehydrogenase complex.
Isocitrate
Fumarate
dehydrogenase
- ATP Can I Keep Selling Sex
- NADH
FADH2
+ ADP For Money, Officer?
α-ketoglutarate
α-KG CO2 + NADH
Succinate dehydrogenase
GTP
+ - Succinyl-CoA
CoA Succinyl-
- NADH
CoA
- ATP

BIOC H E M ISTRY
Electron transport chain and oxidative phosphorylation

ADP + Pi ATP + H2O


NADH NAD+ 1
/2O2 H2O
Oligomycin

CoQ CoQ

Complex I Complex III Complex IV Complex V


2,4-Dinitrophenol
H+ H+ H+ H+

Electron transport 1 NADH → 3 ATP; 1 FADH2 → 2 ATP.


chain
Oxidative 1. Electron transport inhibitors (rotenone, antimycin A, CN−, CO) directly inhibit
phosphorylation electron transport, causing a ↓ of proton gradient and block of ATP synthesis.
poisons 2. ATPase inhibitor (oligomycin) directly inhibits mitochondrial ATPase, causing an
↑ of proton gradient, but no ATP is produced because electron transport stops.
3. Uncoupling agents (2,4-DNP) ↑ permeability of membrane, causing a ↓ of
proton gradient and ↑ O2 consumption. ATP synthesis stops. Electron transport
continues.

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Gluconeogenesis, irreversible enzymes


Pyruvate carboxylase In mitochondria. Pyruvate → oxaloacetate. Requires biotin, ATP.
Activated by acetyl-CoA.
PEP carboxykinase In cytosol. Oxaloacetate → phosphoenolpyruvate. Requires GTP.
Fructose-1,6- In cytosol. Fructose-1,6-bisphosphate →
bisphosphatase fructose-6-P.
Pathway Produces
Glucose-6- In cytosol. Glucose-6-P → glucose. Fresh Glucose.
phosphatase
Above enzymes found only in liver, kidney, intestinal epithelium. Muscle cannot
participate in gluconeogenesis.
Hypoglycemia is caused by a deficiency of the key gluconeogenic enzymes listed above
(e.g., von Gierke’s disease, which is caused by a lack of glucose-6-phosphatase in the liver).
BIOC H E M ISTRY

Pentose phosphate Produces ribose-5-P from G6P for nucleotide synthesis.


pathway (HMP Produces NADPH from NADP+ for fatty acid and steroid biosynthesis and for
shunt) maintaining reduced glutathione inside RBCs.
All reactions of this pathway occur in the cytoplasm. No ATP is used or produced.
Sites: lactating mammary glands, liver, adrenal cortex––all sites of fatty acid or steroid
synthesis.

Glucose-6- G6PD is a rate-limiting enzyme in HMP shunt G6PD deficiency is more


phosphate (which yields NADPH). NADPH is necessary prevalent among blacks.
dehydrogenase to keep glutathione reduced, which in turn Heinz bodies––altered
deficiency detoxifies free radicals and peroxides. ↓ NADPH Hemoglobin precipitates
in RBCs leads to hemolytic anemia due to poor within RBCs.
RBC defense against oxidizing agents (fava beans, X-linked recessive disorder.
sulfonamides, primaquine) and antituberculosis
drugs.
G6P NADP+ H2O2
2 GSH
(reduced)

Glucose-6-phosphate Glutathione
dehydrogenase reductase

6PG NADPH GS–SG 2H2O


(oxidized)

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Disorders of fructose metabolism

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Fructose intolerance Hereditary deficiency of aldolase B (recessive). Fructose-1-phosphate accumulates,
causing a ↓ in available phosphate, which results in inhibition of glycogenolysis
and gluconeogenesis.
Symptoms: hypoglycemia, jaundice, cirrhosis, vomiting.
Treatment: must ↓ intake of both fructose and sucrose (glucose + fructose).
Essential fructosuria Involves a defect in fructokinase and is a benign, asymptomatic condition.
Symptoms: fructose appears in blood and urine.

FRUCTOSE METABOLISM (LIVER)

Dihydroxyacetone-P
Fructokinase Aldolase B
Fructose Fructose-1-P Triose Glyceraldehyde-3-P Glycolysis
kinase
ATP Glyceraldehyde
ADP ATP
NADH

BIOC H E M ISTRY
ADP
Deficiency = fructose intolerance NAD
• Deficiency = essential fructosuria Glycerol

Disorders of galactose metabolism


Galactosemia Absence of galactose-1-phosphate uridyltransferase. Autosomal recessive. Damage is
caused by accumulation of toxic substances (including galactitol) rather than absence
of an essential compound.
Symptoms: cataracts, hepatosplenomegaly, mental retardation.
Treatment: exclude galactose and lactose (galactose + glucose) from diet.
Galactokinase Causes galactosemia and galactosuria, galactitol accumulation if galactose is present in diet.
deficiency
GALACTOSE METABOLISM

Galactokinase Uridyl transferase


Galactose Galactose-1-P Glucose-1-P

ATP
ADP UDP-Glu UDP-Gal
Aldose
reductase Glycolysis/
4-epimerase gluconeogenesis
Galactitol

Lactase deficiency Age-dependent and/or hereditary lactose intolerance (blacks, Asians).


Symptoms: bloating, cramps, osmotic diarrhea.
Treatment: avoid milk or add lactase pills to diet.

Essential amino Ketogenic: Leu, Lys. All essential amino acids:


acids Glucogenic/ketogenic: Ile, Phe, Trp. PriVaTe TIM HALL.
Glucogenic: Met, Thr, Val, Arg, His. Arg and His are required
during periods of growth.

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Acidic and basic At body pH (7.4), acidic amino acids Asp and Glu Asp = aspartic ACID, Glu =
amino acids are negatively charged; basic amino acids Arg glutamic ACID.
and Lys are positively charged. Basic amino Arg and Lys have an extra
acid His at pH 7.4 has no net charge. NH3 group.
Arginine is the most basic amino acid. Arg and Lys
are found in high amounts in histones, which
bind to negatively charged DNA.

Transport of ammonium by alanine and glutamine

Amino acids α-ketoglutarate Alanine Alanine α-ketoglutarate


(NH3) (NH3) (NH3)

α-ketoacids Glutamate Pyruvate Pyruvate Glutamate (NH3)


(NH3)
BIOC H E M ISTRY

Glucose Glucose Urea (NH3)

Muscle Liver

NAD(P)+ NAD(P)H

Glutamine Glutamate α-ketoglutarate

NH4 + NH4+

Urea cycle Degrades amino acids into amino groups. Accounts Ordinarily, Careless Crappers
for 90% of nitrogen in urine. Urea cycle occurs Are Also Frivolous About
in the liver; carbamoyl phosphate incorporation Urination.
occurs in the mitochondria; the remaining steps
occur in the cytosol.
CO2 + NH4+

Citrulline Aspartate
Carbamoyl
phosphate

Mitochondria
(Liver)
Cytoplasm Ornithine Argininosuccinate

Urea

H2O Arginine Fumarate

94
Amino acid derivatives Thyroxine Melanin

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Phenylalanine Tyrosine Dopa Dopamine NE Epi

Niacin NAD+/NADP+
Tryptophan Serotonin
Melatonin

Histidine Histamine

Glycine Porphyrin Heme

Arginine Creatine
Urea
Nitric oxide
Glutamate GABA

BIOC H E M ISTRY
Phenylketonuria Normally, phenylalanine is converted into tyrosine Screened for at birth.
(nonessential aa). In PKU, there is ↓ phenylalanine Phenylketones––phenylacetate,
Phenylalanine
Tyrosine hydroxylase or ↓ tetrahydrobiopterin cofactor. phenyllactate, and
Phenylalanine Tyrosine becomes essential and phenylalanine phenylpyruvate.
hydroxylase
builds up, leading to excess phenylketones in urine. Autosomal-recessive disease.
THB DHB Findings: mental retardation, growth retardation, Incidence ≈ 1:10,000.
fair skin, eczema, musty body odor. Disorder of aromatic amino
NADP+
Dihydropterin
NADPH
Treatment: ↓ phenylalanine (contained in acid metabolism → musty
reductase
aspartame, e.g., NutraSweet) and ↑ tyrosine in diet. body odor.

Alkaptonuria Congenital deficiency of homogentisic acid oxidase in the degradative pathway of


tyrosine. Resulting alkapton bodies cause urine to turn black on standing. Also, the
connective tissue is dark. Benign disease. May have debilitating arthralgias.

Albinism Congenital deficiency of either of the following: Lack of melanin results in an


1. Tyrosinase (inability to synthesize melanin ↑ risk of skin cancer.
from tyrosine)
2. Defective tyrosine transporters (↓ amounts of
tyrosine and thus melanin)
Can result from a lack of migration of neural
crest cells.

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Homocystinuria 3 forms: Results in excess homocysteine


1. Cystathionine synthase deficiency (treatment: in the urine. Cysteine
↓ Met and ↑ Cys in diet) becomes essential.
2. ↓ affinity of cystathionine synthase for Can cause mental retardation,
pyridoxal phosphate (treatment: ↑↑ vitamin osteoporosis, tall stature,
B6 in diet) kyphosis, lens subluxation
3. Methionine synthase deficiency (downward and inward), and
CH3 atherosclerosis (stroke and
via SAM
Cystathionine MI).
synthase
Methionine Methionine Homocysteine Cystathionine Cysteine
synthase B6

B12
THF CH3 THF
BIOC H E M ISTRY

Cystinuria Common (1:7000) inherited defect of renal tubular COLA.


amino acid transporter for Cystine, Ornithine, Treat with acetazolamide to
Lysine, and Arginine in kidneys. Excess cystine alkalinize the urine.
in urine can lead to the precipitation of cystine
kidney stones.

Maple syrup urine Blocked degradation of branched amino acids Urine smells like maple syrup.
disease (Ile, Val, Leu) due to ↓ α-ketoacid dehydrogenase. I Love Vermont maple syrup.
Causes ↑ α-ketoacids in the blood, especially Leu.
Causes severe CNS defects, mental retardation, and
death.

96
Purine salvage deficiencies

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Nucleic acids Nucleic acids

Guanylic acid Inosinic acid Adenylic acid


(GMP) (IMP) (AMP)

3
1 Guanosine 1 Inosine Adenosine
2

Guanine Hypoxanthine
Adenine
4

1 HGPRT + PRPP
Xanthine
4 2 APRT + PRPP
3 Adenosine deaminase (ADA)
Uric acid

BIOC H E M ISTRY
4 Xanthine oxidase

Adenosine ADA deficiency can cause SCID. Excess ATP and SCID––severe combined
deaminase dATP imbalances nucleotide pool via feedback (T and B) immunodeficiency
deficiency inhibition of ribonucleotide reductase. This disease. SCID happens to
prevents DNA synthesis and thus ↓ lymphocyte kids (remember “bubble
count. 1st disease to be treated by experimental boy”).
human gene therapy.
Lesch-Nyhan Purine salvage problem owing to absence of LNS––Lacks Nucleotide
syndrome HGPRTase, which converts hypoxanthine to Salvage (purine).
inosine monophosphate (IMP) and guanine to
guanosine monophosphate (GMP). X-linked
recessive. Results in excess uric acid production.
Findings: retardation, self-mutilation, aggression,
hyperuricemia, gout, and choreoathetosis.

Liver: fed state vs. Digestive system


fasting state
Amino acids
Glucose Amino Chylomicrons glycerol
lactate Fatty
acids acids

Glu AA CM Glycogen TCA cycle

Glu-6-P Fatty
Protein
acids
G6P Pyruvate Acetyl-CoA
HMP
Glycolysis /TCA Fats
shunt
cycle
Glu
Glycogen Ketone
VLDL bodies

FED STATE FASTING STATE


Glucose
In the PHasting state,
PHosphorylate.

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Insulin Made in β cells of pancreas. Required for adipose Insulin moves glucose Into cells.
and skeletal muscle uptake of glucose. BRICK L (don’t need insulin
C peptide GLUT2 receptors are found in β cells and GLUT4 for glucose uptake):
in muscle and fat. Insulin inhibits glucagon Brain
S S
-COOH
Cys
release by α cells of pancreas. RBCs
Cys Cys
Serum C-peptide is not present with exogenous Intestine
Cys S
NH2- S α chain
S S
Cys
insulin intake. Cornea
Cys
β chain
Human proinsulin
Anabolic effects of insulin: Kidney
1. ↑ glucose transport Liver
2. ↑ glycogen synthesis and storage
3. ↑ triglyceride synthesis and storage
4. ↑ Na retention (kidneys)
5. ↑ protein synthesis (muscles)
BIOC H E M ISTRY

Insulin vs. Glucagon phosphorylates stuff → turns glycogen synthase OFF and phosphorylase ON.
glucagon Insulin dephosphorylates stuff → turns glycogen synthase ON and phosphorylase OFF.

Glycogen storage 12 types, all resulting in abnormal glycogen metabolism and an accumulation of glycogen
diseases within cells.
Type I Von Gierke’s disease––glucose-6-phosphatase The liver becomes a muscle.
deficiency. (Think about it.)
Findings: severe fasting hypoglycemia, ↑↑ glycogen
in liver, hepatomegaly, ↑ blood lactate.
Type II Pompe’s disease––lysosomal α-1,4-glucosidase Pompe’s trashes the Pump
deficiency. (heart, liver, and muscle).
Findings: cardiomegaly and systemic findings, leading
to early death.
Type III Cori’s disease––deficiency of debranching enzyme
α-1,6-glucosidase.
Findings: milder form of type I with normal blood
lactate levels.
Type V McArdle’s disease––skeletal muscle glycogen McArdle’s: Muscle.
phosphorylase deficiency.
Findings: ↑ glycogen in muscle but cannot break it Very Poor Carbohydrate
down, leading to painful cramps, myoglobinuria Metabolism.
with strenuous exercise.

98
Lysosomal storage Each is caused by a deficiency in one of the many lysosomal enzymes.

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diseases
Accumulated
Disease Findings Deficient enzyme substrate Inheritance
Fabry’s disease Peripheral neuropathy of α-galactosidase A Ceramide XR
hands/feet, angiokeratomas, trihexoside
cardiovascular/renal disease
Gaucher’s disease Hepatosplenomegaly, β-glucocerebrosidase Glucocerebroside AR
aseptic necrosis of femur,
bone crises, Gaucher’s cells
(macrophages)
Niemann-Pick Progressive neurodegeneration, Sphingomyelinase Sphingomyelin AR
disease hepatosplenomegaly, cherry-
red spot (on macula)
Tay-Sachs disease Progressive neurodegeneration, Hexosaminidase A GM2 ganglioside AR
developmental delay,

BIOC H E M ISTRY
cherry-red spot, lysozymes
with onion skin
Krabbe’s disease Peripheral neuropathy, β-galactosidase Galactocerebroside AR
developmental delay,
optic atrophy
Metachromatic Central and peripheral Arylsulfatase A Cerebroside sulfate AR
leukodystrophy demyelination with ataxia,
dementia
Hurler’s syndrome Developmental delay, α-L-iduronidase Heparan sulfate, AR
gargoylism, airway dermatan sulfate
obstruction, corneal
clouding,
hepatosplenomegaly
Hunter’s syndrome Mild Hurler’s + aggressive Iduronate sulfatase Heparan sulfate, XR
behavior, no corneal dermatan sulfate
clouding
GM1
No man picks (Niemann-Pick)
his nose with his sphinger
GM2
Globoside (sphingomyelinase).
Tay-Sachs
Tay-SaX (Tay-Sachs)
Ceramide trihexoside lacks heXosaminidase.
GM3
Hunters aim for the X
Fabry´s
(X-linked recessive).
Sulfatides Lactosyl cerebroside
Metachromatic
leukodystrophy
Glucocerebroside
Galactocerebroside
Gaucher's
Sphingomyelin
Krabbe´s
Cerebroside
Niemann-Pick

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Fatty acid
metabolism sites
Fatty acid synthesis

Malonyl-CoA

CO2 (biotin) Fatty acid + CoA


Acetyl-CoA
Fatty acid CoA
synthetase

Acyl-CoA
- Malonyl-CoA
Cell cytoplasm
Inner mitochondrial Citrate Carnitine
membrane shuttle shuttle
Mitochondrial matrix
BIOC H E M ISTRY

Acetyl-CoA Acyl-CoA Fatty acid degradation


occurs where its
β-oxidation products will be
(breakdown to consumed—in the
acetyl-CoA groups) mitochondrion.

Ketone bodies In liver: fatty acid and amino acids → acetoacetate + Breath smells like acetone
β-hydroxybutyrate (to be used in muscle and brain). (fruity odor). Urine test for
Ketone bodies found in prolonged starvation and ketones does not detect
diabetic ketoacidosis. Excreted in urine. Made from β-hydroxybutyrate (favored
HMG-CoA. Ketone bodies are metabolized by the by high redox state).
brain to 2 molecules of acetyl-CoA.

Cholesterol Rate-limiting step is catalyzed by HMG-CoA Lovastatin inhibits HMG-


synthesis reductase, which converts HMG-CoA to CoA reductase.
mevalonate. 2⁄3 of plasma cholesterol is esterified
by lecithin-cholesterol acyltransferase (LCAT).

100
Lipoproteins

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C-II
Lipoprotein
Less lipase TG
TG B-100 B-100
CE
CE
VLDL
Hepatic IDL E E
triglyceride
lipase

CE B-100 Receptor
for B-100 E
TG
LDL FFA
B-4
8

Chylomicron
remnant
100

48
B- Lipoprotein

B-
CE lipase

BIOC H E M ISTRY
TG A
Modified Eat TG
LDL
E Chylomicron

C-
II
Pancreatic
lipase Intestinal cells convert FFA back to TG
and package in chylomicrons
Atherosclerotic
plaque
FFA
Small intestine

TG = triglyceride, CE = cholesterol, FFA = free fatty acid

Pancreatic lipase––degradation of dietary TG in small intestine.


Lipoprotein lipase––degradation of TG circulating in chylomicrons and VLDLs.
Hepatic TG lipase––degradation of TG remaining in IDL.
Hormone-sensitive lipase––degradation of TG stored in adipocytes.

Major A-I––Activates LCAT.


apolipoproteins B-100––Binds to LDL receptor.
C-II––Cofactor for lipoprotein lipase.
E––Mediates Extra (remnant) uptake.

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Lipoprotein functions Lipoproteins are composed of varying proportions of cholesterol, triglycerides,


and phospholipids.
Function and route Apolipoproteins
Chylomicron Delivers dietary triglycerides to peripheral tissues and B-48 mediates secretion.
dietary cholesterol to liver. Secreted by intestinal A’s are used for formation of
epithelial cells. Excess causes pancreatitis, lipemia new HDL.
retinalis, and eruptive xanthomas. C-II activates lipoprotein lipase.
E mediates remnant uptake by
liver.
VLDL Delivers hepatic triglycerides to peripheral tissues. B-100 mediates secretion.
Secreted by liver. Excess causes pancreatitis. C-II activates lipoprotein lipase.
E mediates remnant uptake by
liver.
BIOC H E M ISTRY

IDL Formed in the degradation of VLDL. Delivers


triglycerides and cholesterol to liver, where they
are degraded to LDL.
LDL Delivers hepatic cholesterol to peripheral tissues. B-100 mediates binding to cell
Formed by lipoprotein lipase modification of surface receptor for
VLDL in the peripheral tissue. Taken up by target endocytosis.
cells via receptor-mediated endocytosis. Excess
causes atherosclerosis, xanthomas, and arcus
corneae.
HDL Mediates centripetal transport of cholesterol (reverse A’s help form HDL structure.
cholesterol transport, from periphery to liver). Acts A-I in particular activates
as a repository for apoC and apoE (which are LCAT (which catalyzes
needed for chylomicron and VLDL metabolism). esterification of cholesterol).
Secreted from both liver and intestine. CETP mediates transfer of
cholesteryl esters to other
lipoprotein particles.
LDL and HDL carry most cholesterol. LDL HDL is Healthy.
transportscholesterol from liver to tissue; HDL LDL is Lousy.
transports it from periphery to liver.

Familial dyslipidemias

Elevated
Type Increased blood levels Pathophysiology
I––hyperchylomicronemia Chylomicrons TG, cholesterol Lipoprotein lipase deficiency
or altered apolipoprotein C-II
IIa––hypercholesterolemia LDL Cholesterol ↓ LDL receptors
IIb––combined hyperlipidemia LDL, VLDL TG, cholesterol Hepatic overproduction of VLDL
III––dysbetalipoproteinemia IDL, VLDL TG, cholesterol Altered apolipoprotein E
IV––hypertriglyceridemia VLDL TG Hepatic overproduction of VLDL
V––mixed hypertriglyceridemia VLDL, chylomicrons TG, cholesterol ↑ production/↓ clearance of
VLDL and chylomicrons

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Heme synthesis Underproduction of heme causes microcytic hypochromic anemia. Accumulation of

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intermediates causes porphyrias.

Succinyl CoA + Glycine Heme


Committed step
(−) ALA synthetase
Lead Fe2
δ-Aminolevolinic acid poisoning Protoporphyrin
(ALA)

Porphobilinogen Coproporphyrinogen
Acute intermittent Porphyria
porphyria cutanea tarda
Pre-uroporphyrinogen Uroporphyrinogen III

Porphyrias
Lead poisoning Inhibits ferrochelatase and ALA dehydrase.

BIOC H E M ISTRY
Coproporphyrin and ALA accumulate in urine.
Acute intermittent Deficiency in uroporphyrinogen I synthetase. Symptoms = 5 P’s: Painful
porphyria Porphobilinogen and δ-ALA accumulate in urine. abdomen, Pink urine,
Porphyria cutanea Deficiency in uroporphyrinogen decarboxylase. Polyneuropathy,
tarda Uroporphyrin accumulates in urine (tea-colored). Psychological disturbances,
Photosensitivity. Precipitated by drugs.

Heme catabolism Heme is scavenged from RBCs and Fe2+ is reused. Heme → biliverdin → bilirubin (sparingly
water soluble, toxic to CNS, transported by albumin). Bilirubin is removed from blood by
liver, conjugated with glucuronate, and excreted in bile. In the intestine it is processed
into its excreted form. Some urobilinogen, an intestinal intermediate, is reabsorbed into
blood and excreted as urobilin into urine.

Hemoglobin Hemoglobin is composed of 4 polypeptide subunits Carbon monoxide has 200×


β1
(2 α and 2 β) and exists in 2 forms: greater affinity than O2
β2
1. T (taut) form has low affinity for O2. for hemoglobin.
2. R (relaxed) form has high affinity for O2
(300×). Hemoglobin exhibits positive
cooperativity and negative allostery (accounts
for the sigmoid-shaped O2 dissociation curve
α2 α1 for hemoglobin), unlike myoglobin.
Heme

Hemoglobin structure ↑ Cl−, H+, CO2, 2,3-BPG, and temperature favor When you’re Relaxed, you do
regulation T form over R form (shifts dissociation curve to your job better (carry O2).
right, leading to ↑ O2 unloading). Fetal hemoglobin (2α and 2γ
subunits) has lower affinity
for 2,3-BPG than adult
hemoglobin (HbA) and thus
has higher affinity for O2.

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Methemo- Iron in hemoglobin is in a reduced state (ferrous, Administer nitrites in cyanide


globinemia Fe2+). Methemoglobin is an oxidized form of poisoning to oxidize
hemoglobin (ferric, Fe3+) that does not bind O2 hemoglobin to
as readily but has ↑ affinity for CN–. methemoglobin form.
Treat toxic levels of
METHemoglobin with
METHylene blue.

CO2 transport in CO2 binds to amino acids in globin chain (at N CO2 must be transported from
blood terminus) but not to heme. CO2 binding favors T tissue to lungs, the reverse
(taut) form of hemoglobin (and thus promotes O2 of O2 (occurs primarily in the
unloading). form of bicarbonate).
BIOC H E M ISTRY

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Polymerase chain Molecular biology laboratory procedure that is used to synthesize many copies of a desired
reaction (PCR) fragment of DNA.
Steps:
1. DNA is denatured by heating to generate 2 separate strands
2. During cooling, excess premade DNA primers anneal to a specific sequence on each
strand to be amplified
3. Heat-stable DNA polymerase replicates the DNA sequence following each primer
These steps are repeated multiple times for DNA sequence amplification.

Molecular biology techniques


Southern blot A DNA sample is electrophoresed on a gel and then SNoW DRoP:
transferred to a filter. The filter is then soaked in a Southern = DNA
denaturant and subsequently exposed to a labeled Northern = RNA
DNA probe that recognizes and anneals to its Western = Protein

BIOC H E M ISTRY
complementary strand. The resulting double-
stranded labeled piece of DNA is visualized
when the filter is exposed to film.
Northern blot Similar technique, except that Northern blotting
involves radioactive DNA probe binding to
sample RNA.
Western blot Sample protein is separated via gel electrophoresis
and transferred to a filter. Labeled antibody is
used to bind to relevant protein.

Enzyme-linked A rapid immunologic technique testing for ELISA is used in many


immunosorbent antigen-antibody reactivity. laboratories to determine
assay (ELISA) Patient’s blood sample is probed with either whether a particular
1. Test antigen (coupled to color-generating antibody (e.g., anti-HIV) is
Specific IgG
in patient’s enzyme)––to see if immune system present in a patient’s blood
1. blood recognizes it; or sample. Both the sensitivity
Test 2. Test antibody (coupled to color-generating and the specificity of ELISA
Peroxidase
antigen
enzyme enzyme)––to see if a certain antigen is approach 100%, but both
2. generates present false positive and false
color
If the target substance is present in the sample, negative results do occur.
Test antibody
the test solution will have an intense color
Specific
antigen in
reaction, indicating a positive test result.
patient’s blood

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Genetic terms
Variable expression Nature and severity of the phenotype varies from 1 individual to another.
Incomplete Not all individuals with a mutant genotype show the mutant phenotype.
penetrance
Pleiotropy 1 gene has > 1 effect on an individual’s phenotype.
Imprinting Differences in phenotype depend on whether the mutation is of maternal or paternal
origin (e.g., AngelMan’s syndrome [Maternal], Prader-Willi syndrome [Paternal]).
Anticipation Severity of disease worsens or age of onset of disease is earlier in succeeding generations
(e.g., Huntington’s disease).
Loss of heterozygosity If a patient inherits or develops a mutation in a tumor suppressor gene, the complementary
allele must be deleted/mutated before cancer develops. This is not true of oncogenes.
Dominant negative Exerts a dominant effect. A heterozygote produces a nonfunctional altered protein that
mutation also prevents the normal gene product from functioning.
Linkage Tendency for certain alleles at 2 linked loci to occur together more often than
BIOC H E M ISTRY

disequilibrium expected by chance. Measured in a population, not in a family, and often varies in
different populations.
Mosaicism Occurs when cells in the body have different genetic makeup (e.g., lyonization––
random X inactivation in females).
Locus heterogeneity Mutations at different loci can produce the same phenotype (e.g., albinism).

Hardy-Weinberg If a population is in Hardy-Weinberg equilibrium, Hardy-Weinberg law assumes:


population then: 1. There is no mutation
genetics Disease prevalence: p2 + 2pq + q2 = 1 occurring at the locus
Allele prevalence: p + q = 1 2. There is no selection for
p and q are separate alleles; 2pq = heterozygote any of the genotypes at
prevalence. the locus
3. Mating is completely
random
4. There is no migration into or
out of the population
being considered

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Modes of inheritance

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Autosomal dominant Often due to defects in structural genes. Many Often pleiotropic and, in many
generations, both male and female, affected. cases, present clinically after
puberty. Family history
crucial to diagnosis.

Autosomal recessive 25% of offspring from 2 carrier parents are affected. Commonly more severe than
Often due to enzyme deficiencies. Usually seen in dominant disorders; patients
only 1 generation. often present in childhood.

BIOC H E M ISTRY
X-linked recessive Sons of heterozygous mothers have a 50% chance of Commonly more severe in
being affected. No male-to-male transmission. males. Heterozygous females
carrier
may be affected.

X-linked dominant Transmitted through both parents. Either male or Hypophosphatemic rickets.
female offspring of the affected mother may
be affected, while all female offspring of the
affected father are diseased.

Mitochondrial Transmitted only through mother. All offspring of Leber’s hereditary optic
inheritance affected females may show signs of disease. neuropathy; mitochondrial
myopathies.

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Autosomal-dominant diseases
Adult polycystic kidney Always bilateral, massive enlargement of kidneys due to multiple large cysts. Patients
disease present with pain, hematuria, hypertension, progressive renal failure. 90% of cases
are due to mutation in APKD1 (chromosome 16). Associated with polycystic liver
disease, berry aneurysms, mitral valve prolapse. Juvenile form is recessive.
Familial Elevated LDL owing to defective or absent LDL receptor. Heterozygotes (1:500) have
hypercholesterolemia cholesterol ≈ 300 mg/dL. Homozygotes (very rare) have cholesterol ≈ 700+ mg/dL,
(hyperlipidemia severe atherosclerotic disease early in life, and tendon xanthomas (classically in the
type IIA) Achilles tendon); MI may develop before age 20.
Marfan’s syndrome Fibrillin gene mutation → connective tissue disorders.
Skeletal abnormalities––tall with long extremities (arachnodactyly), hyperextensive
joints, and long, tapering fingers and toes (see Image 109).
Cardiovascular––cystic medial necrosis of aorta → aortic incompetence and
dissecting aortic aneurysms. Floppy mitral valve.
BIOC H E M ISTRY

Ocular––subluxation of lenses.
Neurofibromatosis Findings: café-au-lait spots, neural tumors, Lisch nodules (pigmented iris
type 1 (von hamartomas). Also marked by skeletal disorders (e.g., scoliosis),
Recklinghausen’s pheochromocytoma, and ↑ tumor susceptibility. On long arm of chromosome
disease) 17; 17 letters in von Recklinghausen.
Neurofibromatosis Bilateral acoustic neuroma, optic pathway gliomas, juvenile cataracts. NF2 gene on
type 2 chromosome 22; type 2 = 22.
Tuberous sclerosis Findings: facial lesions (adenoma sebaceum), hypopigmented “ash leaf spots” on skin,
cortical and retinal hamartomas, seizures, mental retardation, renal cysts, cardiac
rhabdomyomas. Incomplete penetrance, variable presentation.
Von Hippel–Lindau Findings: hemangioblastomas of retina/cerebellum/medulla; about half of affected
disease individuals develop multiple bilateral renal cell carcinomas and other tumors.
Associated with deletion of VHL gene (tumor suppressor) on chromosome 3 (3p).
Von Hippel–Lindau = 3 words for chromosome 3.
Huntington’s disease Findings: depression, progressive dementia, choreiform movements, caudate atrophy,
and ↓ levels of GABA and ACh in the brain. Symptoms manifest in affected
individuals between the ages of 20 and 50. Gene located on chromosome 4; triplet
repeat disorder. “Hunting 4 food.”
Familial adenomatous Colon becomes covered with adenomatous polyps after puberty. Progresses to colon
polyposis cancer unless resected. Deletion on chromosome 5; 5 letters in “polyp.”
Hereditary Spheroid erythrocytes; hemolytic anemia; increased MCHC. Splenectomy is curative.
spherocytosis
Achondroplasia Autosomal-dominant cell-signaling defect of fibroblast growth factor (FGF) receptor 3.
Results in dwarfism; short limbs, but head and trunk are normal size.

Autosomal- Cystic fibrosis, albinism, α1-antitrypsin deficiency, phenylketonuria, thalassemias,


recessive sickle cell anemias, glycogen storage diseases, mucopolysaccharidoses (except Hunter’s),
diseases sphingolipidoses (except Fabry’s), infant polycystic kidney disease, hemochromatosis.

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Cystic fibrosis Autosomal-recessive defect in CFTR gene on Infertility in males due to absent

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chromosome 7. Defective Cl− channel → secretion vas deferens. Fat-soluble
of abnormally thick mucus that plugs lungs, vitamin deficiencies (A, D, E,
pancreas, and liver → recurrent pulmonary K). Can present as failure to
infections (Pseudomonas species and S. aureus), thrive in infancy.
chronic bronchitis, bronchiectasis, pancreatic Most common lethal genetic
insufficiency (malabsorption and steatorrhea), disease of Caucasians.
meconium ileus in newborns. ↑ concentration Treatment: N-acetylcysteine
of Cl− ions in sweat test is diagnostic. to loosen mucous plugs.

X-linked recessive Fragile X, Duchenne’s muscular dystrophy, hemophilia A and B, Fabry’s, G6PD
disorders deficiency, Hunter’s syndrome, ocular albinism, Lesch-Nyhan syndrome, Bruton’s
agammaglobulinemia, Wiskott-Aldrich syndrome.
Female carriers of X-linked recessive disorders are rarely affected because of random
inactivation of X chromosomes in each cell.

BIOC H E M ISTRY
Muscular dystrophies
Duchenne’s Frame-shift mutation → deletion of dystrophin Duchenne’s = Deleted
(X-linked) gene → accelerated muscle breakdown. Onset Dystrophin.
before 5 years of age. Weakness begins in pelvic Diagnose muscular dystrophies
girdle muscles and progresses superiorly. by ↑ CPK and muscle
Pseudohypertrophy of calf muscles due to biopsy.
fibrofatty replacement of muscle; cardiac myopathy.
The use of Gowers’ maneuver, requiring assistance
of the upper extremities to stand up, is characteristic
(indicates proximal lower limb weakness).
Becker’s Mutated dystrophin gene is less severe than
Duchenne’s.

Fragile X syndrome X-linked defect affecting the methylation and Triplet repeat disorder (CGG)n
expression of the FMR1 gene. The 2nd most that may show genetic
common cause of genetic mental retardation anticipation (germlike
(the most common cause is Down syndrome). expansion in females).
Associated with macro-orchidism (enlarged testes), Fragile X = eXtra-large
long face with a large jaw, large everted ears, testes, jaw, ears.
and autism.

Trinucleotide repeat Huntington’s disease, myotonic dystrophy, Try (trinucleotide) hunting


expansion diseases Friedreich’s ataxia, fragile X syndrome. May show for my fried eggs (X).
anticipation (disease severity ↑ and age of onset
↓ in successive generations).

Common congenital 1. Heart defects


malformations 2. Hypospadias
3. Cleft lip (with or without cleft palate)
4. Congenital hip dislocation
5. Spina bifida
6. Anencephaly
7. Pyloric stenosis Associated with projectile
vomiting.

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Autosomal trisomies
Down syndrome Most common chromosomal disorder and cause of Drinking age (21).
(trisomy 21), 1:700 congenital mental retardation. Findings: mental ↓ levels of α-fetoprotein,
retardation, flat facial profile, prominent epicanthal ↑ β-hCG, ↑ nuchal
folds, simian crease, duodenal atresia, congenital translucency.
heart disease (most common malformation is
septum primum–type ASD due to endocardial
cushion defects), Alzheimer’s disease in affected
individuals > 35 years old, ↑ risk of ALL.
95% of cases due to meiotic nondisjunction of homologous
chromosomes; associated with advanced maternal
age (from 1:1500 in women < 20 to 1:25 in
women > 45). 4% of cases due to robertsonian
translocation, and 1% of cases due to Down
BIOC H E M ISTRY

mosaicism (no maternal association) (see


Image 110).
Edwards’ syndrome Findings: severe mental retardation, rocker bottom Election age (18).
(trisomy 18), feet, low-set ears, micrognathia (small jaw),
1:8000 congenital heart disease, clenched hands,
prominent occiput. Death usually occurs within
1 year of birth.
Patau’s syndrome Findings: severe mental retardation, microphthalmia, Puberty (13).
(trisomy 13), microcephaly, cleft lip/palate, abnormal forebrain
1:6000 structures, polydactyly, congenital heart disease.
Death usually occurs within 1 year of birth.

NONDISJUNCTION

Anaphase I

Anaphase II

n+1 n+1 n-1 n-1 n-1 n+1 n n

Cri-du-chat Congenital deletion of short arm of chromosome 5 Cri du chat = cry of the cat.
syndrome (46,XX or XY, 5p−).
Findings: microcephaly, severe mental retardation,
high-pitched crying/mewing, epicanthal folds,
cardiac abnormalities.

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22q11 syndromes Cleft palate, Abnormal facies, Thymic aplasia → CATCH-22.

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T-cell deficiency, Cardiac defects, Hypocalcemia
2° to parathyroid aplasia, microdeletion at
chromosome 22q11. Variable presentation as
DiGeorge syndrome (thymic, parathyroid, and
cardiac defects) or velocardiofacial syndrome
(palate, facial, and cardiac defects).

Fetal alcohol Newborns of mothers who consumed significant amounts of alcohol (teratogen) during
syndrome pregnancy (highest risk at 3–8 weeks) have ↑ incidence of congenital abnormalities,
including pre- and postnatal developmental retardation, microcephaly, facial
abnormalities, limb dislocation, and heart and lung fistulas. Mechanism may include
inhibition of cell migration. The number one cause of congenital malformations in
the United States.

BIOC H E M ISTRY

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Vitamins
Vitamins

Fat solub le Water soluble

Vitamin A—Vision Pyridoxine––B6


Vitamin D—Bone calcification Vitamin C Pyridoxal––B6
—Ca 2+ homeostasis Pyridoxamine––B6
Vitamin K—Clotting factors
Vitamin E—Antioxidant Folate––Blood,
Metabolic
–Thiamine––B1 neural development
–Riboflavin––B2 Cobalamin––B12 -blood,
–Niacin––B3 CNS
–Pantothenic acid
–Biotin
BIOC H E M ISTRY

Vitamins: fat A, D, E, K. Absorption dependent on gut (ileum) Malabsorption syndromes


soluble and pancreas. Toxicity more common than (steatorrhea), such as cystic
for water-soluble vitamins, because these fibrosis and sprue, or mineral
accumulate in fat. oil intake can cause fat-
soluble vitamin deficiencies.

Vitamins: water B1 (thiamine: TPP) All wash out easily from body
soluble B2 (riboflavin: FAD, FMN) except B12 (stored in liver).
B3 (niacin: NAD+) B-complex deficiencies often
B5 (pantothenate: CoA) result in dermatitis,
B6 (pyridoxine: PP) glossitis, and diarrhea.
B12 (cobalamin)
C (ascorbic acid)
Biotin
Folate

Vitamin A (retinol)
Deficiency Night blindness, dry skin, and impaired immune Retinol is vitamin A, so think
response. Retin-A (used topically for
Function Constituent of visual pigments (retinal). wrinkles and acne).
Excess Arthralgias, fatigue, headaches, skin changes, sore
throat, alopecia.

Vitamin B1 (thiamine)
Deficiency Beriberi and Wernicke-Korsakoff syndrome. Seen Spell beriberi as Ber1Ber1.
in alcoholism and malnutrition. Dry beriberi––polyneuritis,
Function In thiamine pyrophosphate, a cofactor for oxidative muscle wasting.
decarboxylation of α-keto acids (pyruvate, Wet beriberi––high-output
α-ketoglutarate) and a cofactor for cardiac failure (dilated
transketolase in the HMP shunt. cardiomyopathy), edema.

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Vitamin B2 (riboflavin)

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Deficiency Angular stomatitis, Cheilosis, Corneal The 2 C’s.
vascularization. FAD and FMN are derived from
Function Cofactor in oxidation and reduction (e.g., FADH2). riboFlavin (B2 = 2 ATP).

Vitamin B3 (niacin)
Deficiency Pellagra can be caused by Hartnup disease Pellagra’s symptoms are the 3
(↓ tryptophan absorption), malignant D’s: Diarrhea, Dermatitis,
carcinoid syndrome (↑ tryptophan Dementia (also beefy
metabolism), and INH (↓ vitamin B6). glossitis).
Function Constituent of NAD+, NADP+ (used in redox NAD derived from Niacin
reactions). Derived from tryptophan (B3 = 3 ATP).
using vitamin B6.

Vitamin B5 (pantothenate)
Deficiency Dermatitis, enteritis, alopecia, adrenal insufficiency.

BIOC H E M ISTRY
Function Constituent of CoA (a cofactor for acyl transfers) Pantothen-A is in Co-A.
and component of fatty acid synthase.

Vitamin B6 (pyridoxine)
Deficiency Convulsions, hyperirritability (deficiency inducible by INH and oral contraceptives),
peripheral neuropathy.
Function Converted to pyridoxal phosphate, a cofactor used in transamination (e.g., ALT and AST),
decarboxylation, and heme synthesis.

Vitamin B12 (cobalamin)


Deficiency Macrocytic, megaloblastic anemia; neurologic Found only in animal products.
symptoms (optic neuropathy, subacute combined Vitamin B12 deficiency is
degeneration, paresthesia); glossitis. usually caused by
Function Cofactor for homocysteine methylation malabsorption (sprue,
(transfers CH3 groups as methylcobalamin) enteritis, Diphyllobothrium
and methylmalonyl-CoA handling. latum), lack of intrinsic factor
Stored primarily in the liver. (pernicious anemia), or
Synthesized only by microorganisms. absence of terminal ileum
(Crohn’s disease).
Use Schilling test to detect
deficiency.
B12 Abnormal myelin is seen in B12
Homocysteine + N-methyl THF Methionine + THF deficiency, possibly due to
B12 ↓ methionine or ↑
Methylmalonyl-CoA Succinyl-CoA methylmalonic acid (from
metabolism of accumulated
methylmalonyl-CoA).

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Folic acid
Deficiency Most common vitamin deficiency in the United FOLate from FOLiage.
States. Eat green leaves (because folic
Macrocytic, megaloblastic anemia (often no acid is not stored very long).
neurologic symptoms, as opposed to vitamin Supplemental folic acid in
B12 deficiency). early pregnancy reduces
Function Coenzyme (tetrahydrofolate) for 1-carbon neural tube defects.
transfer; involved in methylation reactions. PABA is the folic acid
Important for the synthesis of nitrogenous bases precursor in bacteria. Sulfa
in DNA and RNA. drugs and dapsone
(antimicrobials) are PABA
analogs.

Biotin
BIOC H E M ISTRY

Deficiency Dermatitis, enteritis. Caused by antibiotic use, “AVIDin in egg whites


ingestion of raw eggs. AVIDly binds biotin.”
Function Cofactor for carboxylations:
1. Pyruvate → oxaloacetate
2. Acetyl-CoA → malonyl-CoA
3. Proprionyl-CoA → methylmalonyl-CoA

Vitamin C (ascorbic acid)


Deficiency Scurvy––swollen gums, bruising, anemia, poor Vitamin C Cross-links
Function wound healing. Collagen. British sailors
Necessary for hydroxylation of proline and lysine in carried limes to prevent
collagen synthesis. scurvy (origin of the word
Facilitates iron absorption by keeping iron in Fe+2 “limey”).
reduced state (more absorbable)
Necessary as a cofactor for dopamine → NE.

Vitamin D D2 = ergocalciferol, consumed in milk. Remember that drinking milk


D3 = cholecalciferol, formed in sun-exposed skin. (fortified with vitamin D) is
25-OH D3 = storage form. good for bones.
1,25 (OH)2 D3 = active form.
Deficiency Rickets in children (bending bones), osteomalacia
in adults (soft bones), and hypocalcemic tetany.
Function ↑ intestinal absorption of calcium and phosphate.
Excess Hypercalcemia, loss of appetite, stupor. Seen in
sarcoidosis, a disease where the epithelioid
macrophages convert vitamin D into its active form.

Vitamin E
Deficiency Increased fragility of erythrocytes. Vitamin E is for Erythrocytes.
Function Antioxidant (protects erythrocytes from hemolysis).

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Vitamin K

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Deficiency Neonatal hemorrhage with ↑ PT and ↑ aPTT but K for Koagulation. Note that
normal bleeding time. the vitamin K–dependent
Function Catalyzes γ-carboxylation of glutamic acid residues clotting factors are II, VII,
on various proteins concerned with blood clotting. IX, X, and protein C and S.
Synthesized by intestinal flora. Therefore, vitamin Warfarin is a vitamin K
K deficiency can occur after the prolonged use of antagonist.
broad-spectrum antibiotics.

Zinc deficiency Delayed wound healing, hypogonadism, ↓ adult hair (axillary, facial, pubic); may
predispose to alcoholic cirrhosis.

Ethanol metabolism Disulfiram (Antabuse) inhibits


Alcohol Acetaldehyde
dehydrogenase dehydrogenase acetaldehyde dehydrogenase
Ethanol Acetaldehyde Acetate (acetaldehyde accumulates,
NAD+ NADH NAD+ NADH contributing to hangover

BIOC H E M ISTRY
symptoms).
NAD+ is the limiting reagent.
Alcohol dehydrogenase operates via zero-order
` kinetics.

Ethanol Ethanol metabolism ↑ NADH/NAD+ ratio in liver, causing diversion of pyruvate


hypoglycemia to lactate and OAA to malate, thereby inhibiting gluconeogenesis and leading to
hypoglycemia. This altered NADH/NAD+ ratio is responsible for the hepatic fatty
change (hepatocellular steatosis) seen in chronic alcoholics (shunting away from
glycolysis and toward fatty acid synthesis).

NADH NAD+

1. Pyruvate lactate

NADH NAD+

2. Oxaloacetate malate

Kwashiorkor vs. Kwashiorkor––protein malnutrition resulting in skin Kwashiorkor results from a


marasmus lesions, edema, liver malfunction (fatty change). protein-deficient MEAL:
Clinical picture is small child with swollen belly. Malabsorption
Marasmus––protein-calorie malnutrition resulting in Edema
tissue wasting. Anemia
Liver (fatty)

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