Proteins

1) Types
1. Catalysts (Enzymes)
a) DNA Polymerase
b) Catalase
c) CK2 Kinase

2. Transport & Storage

a) Hemoglobin
b) Serum albumin
c) Ion channels
d) Ovalbumin

3. Structural
a) Collagen
b) Keratin
c) Silk Fibroin

4. Generate Movement
a) Actin
b) Myosin

5. Regulation of Metabolism and Gene Expression

a) Insulin
b) Lac repressor

6. Protection
a) Immunoglobulin
b) Thrombin and Fibrinogen
c) Venom Proteins

2) Amino Acids (AA) are the building blocks of peptides and

proteins

Peptides generally contain 2-10 AA

Polypeptides contain 10-100 AA
Proteins contain >100 AA

3) L-Amino Acids are the biologically relevant enantiomer

4) The 20 AA can be grouped according to functional classes

Aliphatic (GAVLIMP)
Aromatic (WYF)
Polar, uncharged (CHNQST)
Polar, charged acidic (DE)
 Polar, charged basic (RK)

5) Isoelectric point (pI) of an AA (or peptide/protein) is equal to

the pH when net charge = 0
AA net charge is (-) if pH > pI,
and (+) if pH < pI

6) Primary Structure refers to AA sequence in the protein

Conventionally written from N to C terminus
7) AA can link together via AMIDE BOND to form peptides
Two ends are form: amino or N terminus and carboxyl or C
terminus
Peptide formation is a condensation reaction (loss of H2O)
Water molecule is eliminated in a bond
Peptide bonds can be broken or hydrolyzed by exo- or endo
peptidases
Remaning portions of amino acids are called Residues

8) The peptide bond is planar

9) Note that not all phi (Amino bond to alpha carbon) and psi
(Carboxyl bond to alpha carbon) values are possible

10) Peptides are cleaved via hydrolysis

Acids, bases or enzymes can be used to facilitate the hydrolysis
In our stomach or intestine, peptidases or proteases are present
Enzymes specific to some AA are used for protein analysis
PROTEIN HYDROLYSIS

11) Protein folds naturally to form its 2o and 3o structure

Chaperones and Chaperonins: aid in proper protein folding
(video)

12) Phi and Psi values (recall: 1o)affect the secondary structure
of proteins
helix
sheets (or strands)
turns (or loops)

13) Alpha-helix has 3.6 residues per turn [i.e., AA1-AA2 = 100o]
rodlike structure
Sidechains extend outward in a helical array
stabilized by H-bonds between the NH and CO groups of the
main group
 In a helix the 1st and 19th are exactly on the same side
(1800 degrees)

14) Alpha helices are disrupted by AA that cannot conform to

its structure
Proline / hydroxyproline
amino acids with bulky R groups
presence of charged amino acids

15) sheets are formed by linking 2 or more strands by H-

bonding
sheets may be parallel or anti-parallel
R groups of amino acids lie above or below the plane
16) turns connect the beta sheets
globular shape of proteins is due to the reversal of the direction
of their polypeptide chain
17) Supersecondary structures (motifs) may also be formed

18) Helices will form except:

WTF IVY: -sheet (steric effects)
NDPSG: -turns (H-bonding competition, G = small R
group, P = turn is sharp)
19) Gibbs Free Energy (G) is a measure of stability. Negative
G = spontaneous!

20) The native form of the protein is the most stable form

Protein folding is driven by hydrophobic effect: entropy-

driven

21) S-S bond also affects the tertiary structure of proteins

22) The overall 3D structure is described by its tertiary

structure

23) Quaternary structures show interactions between other

subunits

Hemoglobin: tetramer (two homodimers) with Fe prosthetic

group in each subunit.

24) Heme group of hemoglobin is supported by the tertiary

structure

25) Biologically active proteins are in their native tertiary

structure

26) Proteins when subjected to some conditions are denatured

Heat breaking IMF

pH changing +/- to neutral and vv

Detergents polar and nonpolar groups

Reducing agents (DTT) disulfide bonds

SKELETAL AND MOTOR PROTEINS

1) Proteins can also be classified as fibrous or globular

Globular water soluble, usually biological roles

Fibrous water insoluble, usually structural roles

fibrous proteins generally have only primary and secondary structure whereas globularproteins
have tertiary and sometimes quaternary structure in addition to primary and secondary structure.

2) FIBROUS PROTEINS
play structural roles in animal cells and tissues
Include major proteins of skin and connective tissue
Low solubility in water
3) KERATINS
and keratins
Members of a broad group of intermediate filament

proteins
 4) COLLAGEN
most abundant single protein in most vertebrates
form the matrix material in bone, major portions of

tendons, important constituent of the skin

tropocollagen basic unit
Individual chains are left hand helices
3 of these helices wrap around each other in a right hand

sense

Collagen Composition

every third residue is Gly

Gly Pro Y and Gly-X- HyP

in order to properly serve the multiple functions it does,

collagen exist in a large number of genetic variants

5) Scurvy is caused by lack of Vitamin C

COLLAGEN SYNTHESIS

collagen undergoes extensive modification

post translational modification

starts in the endoplasmic reticulum then to the cytosol and

finally into the extracellular space

6) ELASTIN
found in ligaments and arterial blood vessels
rich in glycine, alanine and valine
conformation approximates that of a random coil
Also contains lysine which can be involved in cross-linking
Lysyl amino oxidase specific to: Lys-Ala-Ala-Lys or Lys-Ala-

Ala-Ala-Lys

o It cross-links the tropocollagen units to form

covalently bundled, cross-linked helices.
 7) MOTOR PROTEINS
Converts chemical energy (ATP) to mechanical energy

(movement)

8) ACTIN-MYOSIN

Muscle contraction [not limited]

Ca2+ conformational change in tropomyosin myosin-binding

site in actin exposed myosin can move actin

9) ACTIN

Responsible for shape and movement of the cell (cell crawling)

aside from muscle contraction

Microfilaments F-actin formed from G-actin units

polymerizes / depolymerizes based on actin monomer

concentration driven by hydrolysis of ATP

(+) end polymerizes faster than (-) end [the ATP-binding end]

10) MYOSIN

Tail = make up the thick filaments in the sacromere

Head = pulls actin fibers together

Light chains make the neck rigid

Hydrolysis of ATP in myosin head induces conformational change

in the actin-binding site. (3.5nm away:

 High affinity for actin: myosin andmyosin-ADP + Pi ;

low affinity for actin: myosin-ATP

11) KINESIN-MICROTUBULE

Involves in mitosis, meiosis, and intracellular vesicle transport

 Mostly involved in center to peripheral movement

12) MICROTUBULE

Alpha monomer and Beta Monomer

ABABABABAB Protofilament
Actin- Need ATP Tubulin= Need GTP

-tubulin and -tubulin dimers form hollow microtubule therefore

much more rigid (microfilaments < intermediate filaments <

microtubules)

Each tubulin has a GTP-binding site

(+) end and (-) end too

13) KINESIN

Two heads (for microtubules) and two tails (for cargo load)

Head has two binding sites: ATP and tubulin

2 ATPs needed to walk 16nm

Neck is elastic to allow walking (not rigid like myosin cannot

swing)

1) GLOBULAR PROTEINS

polypeptide chains folded into compact structures

3-D structure

often has pockets that contains prosthetic groups

each conformation is suited to a particular functional role that

the protein plays

may contain one or more domains

2) General Rules in Tertiary Folding

 1. All globular proteins have defined conformations inside and

outside

2. -sheets are usually twisted or wrapped into barrel structures

3. The polypeptide chain can turn corners in a number of ways

4. Not all parts of a protein can be conveniently classified as helix,

-sheet, or turn

5. the polypeptide sequence determines the proper folding of a

globular protein

6. the protein knows its own favored conformation

7. Only in stability can it function

Hemoglobin and Myoglobin

Binding of oxygen to haemoglobin is known as co-operative binding

because the binding of successive O2 molecules facilitates binding of
the next. Binding of the 1st O2 molecule increases the affinity of
haemoglobin for oxygen and hence facilitates the binding of the 2nd
O2 molelcule
Relaxed (R) state - this state corresponds to the quaternary
strucure of oxyhemoglobin & favours oxygen binding

Tense (T) state - this state corresponds to the quaternary

structure of deoxyhemoglobin & has a lower binding affinity for
oxygen

Transition from one state to another involves the breaking or

formation of salt bridges between the polypeptide chains.
2,3-Diphosphoglycerate:
This molecule binds specifically to the deoxygenated form of
haemoglobin and not the oxygenated form enabling the
unloading of oxygen at the tissues.
1) Oxygen-binding proteins

O2 is nonpolar, but we need them

 Aerobic processes for energy = all processes need O2

heart skip a beat

Myoglobin / Hemoglobin has a heme group (with Fe2+)

If Fe2+ Fe3+, oxygen cannot bind (Metmyoglobin = brownish)

2) Heme group

Heme group is where O2 binds.

Why do we need all the 141 AA of hemoglobin unit / 153AA of

myoglobin (tertiary structure)?

Heme group alone: O2 oxidizes Fe to Fe3+. (protein = protection)

Heme group alone: can bind to CO and O2 (CO 25,000x more)

BUT! Histidine groups modulate this. (CO binds 250x more

instead)

3) Conformational change
 Binding of O2 causes heme group to be planar. (0.029 nm

increase)

Not much effect on myoglobin, but very important for

hemoglobin!

4) Myoglobin vs Hemoglobin

Myoglobin found in muscle, storage of O2

Hemoglobin blood stream, transport of O2
Release and uptake of Hb is more regulated!
pO2 = 100 Torr (lungs) uptake!
pO2 = 40 Torr (capillaries) release!
5) Allosteric effects
Hemoglobin has quaternary structure 22
Homotropic= Amt of oxygen that binds to hemoglobin.

OXYGEN
Heterotropic something else that binds to Hemoglobin

that makes it want to bind to Oxygen. (H+, BPG)

Each monomeric unit ~ myoglobin
subunit is contact [more] with subunits
Contacts are altered when O2 binds to heme
15o conformational change allows binding to other 3

subunits.
Allosteric effects
T state = deoxyHb; R state = oxyHb
O2 can only bind to units if R state [sterically-hindered]

6) Bohr Effect

Amount of Oxygen needed to let hemoglobin know where it will

go.
High H+ -> Release Oxygen
Carbon dioxide makes blood acidic, hence releases Oxygen

Other allosteric factor: H+ and CO2 improves release of O2

(muscles)
 Lower pH protonates His146 so it can form salt bridge with Asp94

CO2 + H2O H2CO3 H+ + HCO3-

7) 2,3-bisphosphoglycerol control

2,3-BPG binds to Hb inc release of O2

High altitude-> Low Oxygen -> Release Bpg to bind to

hemoglobin to release Oxygen

Bpg binds when hemoglobin is closed

1 BPG= 1 Hemoglobin

BPG can fit in -chains in deoxy; too small for oxy-Hb

8) Fetal hemoglobin binds to O2 better

Fetal Hb (HbF) = low affinity for 2,3-BPG, so binds to O2

better!

9) Sickle Cell Anemia

Variations in Hb causes several diseases

HbS: E6V mutation causes a Val sidechain that fits in a pocket

in another b-unit polymerization

HbS is less soluble in water

Immunology

The study of molecules, cells, organs & systems responsible for

RECOGNITION & DISPOSAL of foreign or non-self materials

It deals with understanding the DESIRABLE (Protective) and the

UNDESIRABLE (Pathologic) consequences of immune interactions

1) The Immune System

Primary lymphoid organs

Provide the microenvironments for lymphocyte maturation
Secondary lymphoid organs
Trap antigen from tissues or vascular spaces and provide sites
where mature lymphocytes can interact effectively with the
antigen

2) Innate Immunity
Consits of cellular and biochemical defense mechanisms that are
in place even before infection
React only to microbes and to non-infectious substances
Principal components: physical and chemical barriers,
phagocytic cells and NK cells, blood proteins (complement
system), and cytokines
Phagocytic cells

3) Injury to tissue will cause release of inflammatory chemicals

(i.e. histamine) which will dilate capilliaries to allow
phagocytes to squeeze through DIAPEDESIS

3.5) Phagocytic cell- creates vesicle to engulf cell, diff. granulocytes kill
it differently. Lysosome fuses w/phago cell->Phagolysosome brings
it to B and T cells.

4) Natural killer Cells (NK Cells)

1st- MHC1 is an inhibitor of cytotoxic granule release to kill cell
2nd- MHC1 is not binded, causing cytotoxic release
3rd- MHC1 is binded but high expression of ligands for activation
receptor allows activation to occur.
4.2) P- Selectin-> Produced during inflammation wherein neutrophils
binds to it.
4.3) Chemokines ->Family of Cytokines-> Actin-> Allow movement
for leukocytes->more chemokines, more leukocytes go there.
4.4) Leukocytes move in a concentration gradient.
4.5) Cytokines= Small molecules that encourages growth of
leukocytes. Replication of Leukocytes( White blood cells).

5) Complement System
Composed of more than 12 types of plasma proteins
these proteins act in various combinations to coat the cell and
promote phagocytosis (opsonization), make holes in the cell
membrane (cytolysis), attract leukocytes to the area
(chemotaxis), and otherwise enhance the inflammatory
response.

6) Adaptive Immunity
2 types: cellular and humoral immunity
Cellular immunity guards against virally infected cells, fungi,
parasites and foreign tissue. Mediated by T-cells
 Humoral immunity most effective against bacterial infection
and extracellular phases of viral infection. Mediated by B-cells
(through production of antibodies

Innate: Granulocytes Usually in Whiteblood cells

-Basophils
-Neutrophils
-Eosinophils
7)Macrophages: Phagocytic cells, engulfs foreign bodies
8) Mast Cells: Mast cells play a key role in the inflammatory process.
When activated, a mast cell rapidly releases its
characteristic granules and various hormonal mediators into the
interstitium
-Histamine dilates blood vessels so fighters can go in.
9) Dendritic Cells- Their main function is to process antigen material
and present it on the surface to other cells of the immune system. That
is, dendritic cells function as antigen-presenting cells.

Adaptive Immunity
11) B and T cells =with memory
12) B Cells= Creates antibodies, creates plasma cell.
Plasma Cell- plasmocytes, and effector B cells, are white blood
cells which secrete large volumes of antibodies. They are transported
by the blood plasma and the lymphatic system. Like all blood cells,
plasma cells ultimately originate in the bone marrow

13) T Cells:- CD4- helper cell

CD8- Cytotoxic cell

14) Adaptive immunity- kung hindi siya napatay ni Natural Killer cells.

15) Dynein is a motor protein (also called molecular motor or motor

molecule) in cells which converts the chemical energy contained
inATP into the mechanical energy of movement. Dynein transports
various cellular cargo by "walking"
along cytoskeletalmicrotubules towards the minus-end of the
microtubule, which is usually oriented towards the cell center.
This form of transport is known as retrograde transport. (EX. CILIA)
16) In contrast, kinesins are motor proteins that move toward the
microtubules' plus end, are called plus-end directed motors.

17) MICROFLORA- Good bacteria= compete for space in body for

nutrients. Compete against bad bacteria.

18) Classes of IG= 1st type of IG produced by B cells ->

IgM to fight dengue first ->
Then IgG naman.
IgA- Tears
IgE- Mast Cells
IgD- Di pa sure

CLASSES OF IMMUNOGLOBULINS

19) Epitopes
Places of recognition of antigens (Identify you due to
your eyes, ears, face)
Antigenic Determinants

20) Polyclonal
Recognize different epitopes
Antigen receptors-> Only recognize specific epitopes.

21) Antibody Diversity

 Can create different types of antibodies
Recombinase= tags=shuffle-> To recognize antibodies.

Cellular Immune Response

22) Major Histocompatibility Complex

-MHC molecules mediate interactions of leukocytes

Cellular Immune response

Class 1 MHC-> (Presents to CD8) Cytotoxic to kill
Class 2 MHC-> (Presents to CD4) Helper T Cell to tell B Cell to produce
more antibodies.

23) Macrophages aid in both Cellular and Humoral Immune

Response
Enzyme Kinetics
 A substance that increases the rate, or velocity, of a chemical
reaction without itself being changed in the overall process
Most (but not all) biological catalysts are proteins (abzymes,
ribozymes)
Specific to a substrate
a true catalyst, although it participates in a chemical reaction, is
unchanged by it
Catalysts change the rates of processes but do not affect the
direction/position of equilibrium of the reaction

Enzymes are specific to certain

substrates
1) Active site = where
reaction occurs
Specificity attributed
to the 3D structure of enzymes

2) Hemoglobin composed of hundreds of AA, but important lang ang

2 histidines but you need others to make 2 histidines meet and stable.
3) Restriction Enzymes-> Cleave specific nucleic acid sequences

4) Cofactors -> Needed for enzymes to function

Apoenzyme + Cofactors -> Holoenzyme
No cofactor-> Enzyme is not active
Substrate binds to active site
5) Isozymes -> Different enzymes that catalyze same
reaction (like isomers)
6) Enzymes work best at certain conditions
a. pH and
b. temperature optimization
Denaturation can also occur
Pepsin- PH 2, optimum
Active during acidic conditions in stomach
 If body temp is 38 degrees, that would cause
denaturation of enzymes.
a) Why tertiary structure is changed if pH is changed
Remove and Add hydrogens
It would not fit anymore

7) Two models of fitting are proposed: Lock and Key and Induced Fit

8) Lock and Key

Enzyme perfectly shaped to fit substrate
9) Induced Fit
It induces to fit in substances

8) Mechanisms of catalysis
Acid-base catalysis
Covalent catalysis
Metal ion catalysis

9) HIV-1 protease (acid-base)

10) Serine proteases (Covalent catalysis)

Catalytic triad: Ser, His, Asp (After they do what they do-> They
make peptide go back to the way it was)
Chymotrypsin, trypsin
Scissile bond
11) Serine
Likes to bind to alpha
carbons
Covalent bond
Covalent catalysis
12) Multisubstrate reactions follow either
ping-pong mechanism or sequential

RANDOM: E + A/B EA/EB + B/A EAB E + P

ORDERED: E + A EA + B EAB E + P
 Initial velocity assumes product concentration is almost zero.

KM describes how tight the enzyme binds to the substrate

Tighter-> Faster reaction
Graphically, its [S] when V = Vmax
Substrate concentration at which the reaction velocity is half-
maximal, it indicates how efficiently an enzyme selects its
substrate and converts it to product.
Lower value of KM-> the more effective the enzyme is at low
substrate concentrations
Higher value of KM-> the less effective the enzyme is.
High KM indicates weak binding and low KM indicates strong
binding.

Kcat or turnover number describes the enzyme efficiency

Kcat = Vmax / [E]
13) KM and Aldehyde dehydrogenase
EtOH + NAD+ EtAl + H+ +NADH
Lower Km (mitochondrial ADH) High affinity- mabagal ka
malasing
Higher Km (cytosolic ADH) Low affinity mabilis ka malasing

14) Lineweaver-Burke Graph is an easier way to find KM and Vmax

The enzyme kinetics

can be disrupted by an
INHIBITOR.
3 mechanisms of
inhibition
Competitive
Uncompetitive
Noncompetitive

15) Competitive
inhibitors bind at the
same site with
substrate
Increases KM but
Vmax is the same

16) Noncompetitive
inhibitors bind at a
different site
KM is the same
but Vmax is lowered

17) Uncompetitive inhbitors bind at a different site after substrate

binds
KM and Vmax are changed
 *Active site is usually similar to transition state. Inhibitors
mimic transition state, which is why sometimes the substrate is fooled.

There are also irreversible inhibitions.

18) Formation of breakage of covalent bonds
Ex. DIFP or Aspirin (anti-inflammatory)
-DIFP goes to active site, kills cyclooxygenase that produces
inflammation

19) Allosteric enzymes are governed by effector molecules

Effectors = molecular stoplight
Effector binds to regulatory subunit, to accept (positive effector)
or not accept (negative effector).
Substrate binds more or less depending on whether the effector
is positive or negative.

20) Intermolecular forces reversible

Covalent forces- irreversible

21) Allosteric control can also be covalent

Zymogens = precursor of enzymes
Zymogen (Inactive) --- Active Enzyme
Collagen Procollagen
Insulin Proinsulin
Trypsin Trypsinogen

PH makes lysozymes active, lower ph makes it active. Not too much

lysosomes since it might digest cells.