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NUCLEIC ACIDS I.

GENERAL DESCRIPTION Polymeric molecules in which repeating unit is a nucleotide Discovered by Swiss physiologist, Friedrich Miescher (1844-1895) in 1969 while studying the nuclei of WBCs The fact that they were initially found in the cell nuclei and were acidic accounts for the name nucleic acid II. TYPES OF NUCLEIC ACIDS
Phosphate + Base + Sugar

Phosphate + Sugar = NUCLEOSIDE

Nucleoside + Base = NUCLEOTIDE

FIGURE 1 Components of a Nucleic Acid (DNA)

Backbone of DNA/RNA always the same

VARIABLE: A, U, C, G, T

FIGURE 1A Components of a Nucleic Acid

1. Deoxyribonucleic acid (DNA) Primary function is the storage and transfer of genetic information (is widely used directly to control many functions in the living cells) 2. Ribonucleic acid (RNA) Primarily functions in protein synthesis, the molecules that carry out essential cellular functions

FIGURE 1B Ribonucleotide (RNA) and Deoxyribonucleotide (DNA)

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BASIS Sugar Bases (Pyrimidine) Strands # of nucleotides

RNA

DNA

TABLE 1 Major differences between RNA and DNA

III.

NUCLEOTIDES: Building Blocks of Nucleic Acids Molecule composed of a pentose sugar bonded to both a phosphate group and a nitrogen-containing heterocyclic base

FIGURE 2A Building Blocks of Nucleic Acids

(Fused ring) (Single hetero cyclic ring) FIGURE 2B General Structure of Nitrogen Bases

A.

3 PARTS OF NUCLEOTIDES:

1. Pentose Sugar Sugar unit of nucleotide is either the pentose ribose (RNA) or 2-deoxyribose (DNA) 2. Nitrogen-containing bases/Heterocyclic base i. Purine o Bi-cyclic base with fused five-and-six member rings a. Adenine (A): 6-amino-purine b. Guanine (G): 2-amino-oxypurine ii. Pyrimidine o Monocyclic base with a six-member ring o RNA: Cytosine and Uracil o DNA: Cytosine and Thymine a. Thymine (T): 2,4-dioxy-5-methylpyrimidine b. Cytosine (C): 2-oxy-4-aminopyrimidine c. Uracil (U): 2,4-dioxypyrimidine

3. Phosphate Third component of nucleotide, derived from phosphoric acid (H3PO4) An important nucleotide is adenosine monophosphate (AMP) formed from a reaction of adenosine (a nucleotide) and one molecule of phosphoric acid

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B.

NUCLEOTIDE FORMATION

FIGURE 4 Nucleotide Formation

Important characteristics of this combining of three molecules into one molecule (nucleotide) are: 1. Dehydration (formation of water molecule) occurs at two locations: between the sugar and base, and between the sugar and phosphate. 2. The base is always attached at the C1 position of the sugar. For purine bases, attachment is through N9 for pyrimidine bases, N1 is involved. The C1 carbon atom of the ribose unit is always in configuration. 3. The phosphate group is usually attached to the sugar at C5 position through the phosphate-esterlinkage C. NUCLEOTIDE NOMENCLATURE 1. All of the names end in 5-monophosphate, which signifies the presence of a phosphate group attached to the 5-carbon/atom of ribose or deoxyribose 2. Preceding the monophosphate ending is the name of the base present in the modified form. The suffix osine is used for purine bases, the suffix idine with the pyrimidine bases. 3. The prefix deoxy at the start of the name signifies that the sugar present is deoxyribose, when no prefix is used, the sugar is ribose. 4. The abbreviations in the table for the nucleotides come from the one-letter symbol for the bases (A, C, G, T and U); the use of MP for monophosphate and a lower case d at the start of the abbreviation whenever deoxyribose is the sugar BASE DNA Nucleotides Adenine Guanine Cytosine Thymine RNA Nucleotides Adenine Guanine Cytosine Uracil SUGAR Deoxyribose Deoxyribose Deoxyribose Deoxyribose Ribose Ribose Ribose Ribose NUCLEOTIDE NAME ABBREV.

TABLE 2 Names of Eight Nucleotides found in DNA and RNA

D. PRIMARY STRUCTURE OF NUCLEIC ACIDS Nucleic Acid Backbone o The alternating sugar phosphate chain in nucleic acid structure. o Constant all throughout the entire structure DNA Backbone RNA Backbone
Deoxyribo se Phospha te Phospha te Deoxyribo se Phospha te Phospha te Deoxyribo se

Ribose

Ribose

Ribose Page

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FIGURE 3 Directionality of Nucleic Acid Backbone

PRIMARY STRUCTURE OF A NUCLEIC ACID: The Sequence of Nucleotides in the Molecule 1. Each non-terminal phosphate group of the sugar-phosphate backbone is bonded to two sugar molecules through a 35-phosphodiester linkage. There is a phosphoester bond to the 5 carbon of one sugar unit and a phosphoester bond to the 3 carbon of the other sugar. 2. A nucleotide chain has directionality: one end of the nucleotide chain, the 5 end, normally carries a free phosphate group attached to the 5 carbon atom. The other end of the nucleotide chain, the 3 end, normally has a free hydroxyl group attached to the 3 carbon atom. By convention, the sequence of bases of a nucleic acid strand is read from the 5 end to the 3 end. 3. Each non-terminal phosphate group in the backbone of a nucleic acid carries a -1 charge. The parent phosphoric acid molecule from which the phosphate was derived originally had three OH groups. Two of these become involved in the 3,5-phosphodiester linkage. The remaining OH group is free to exhibit acidic behavior that is, to produce a H+ ion.

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FIGURE 3A Nucleotide Sequence

E.

THE DNA DOUBLE HELIX

The Watson-Crick Model of DNA Proposed a double-coiled consisting of two strands intertwined around one another and held together by hydrogen bonds, much like two entwined rails from spiral staircase The solid lines Dotted lines All DNA molecules have the same sequence of deoxyribose and phosphates in the ladder part of the chain, the difference lies in the order of the adenine, thymine, cytosine and guanine parts of the chain makes up genetic code.

FIGURE 4A Watson-Crick Model of the DNA

FIGURE 4B Hydrogen Bonded Base Pairs

BASE PAIRING Chargaffs Rule o Number of Purine molecules = Number of Pyrimidine molecules A = T G C Complementary Bases o Specific pairs of bases in nucleic acid structures that hydrogen-bond to each other o Two strands of DNA in double helix are complementary. o DNA A C G T o RNA A C G U F. REPLICATION OF DNA MOLECULES:

CHROMOSOMES Individual DNA molecule bound to a group of molecule Prepared by: Sunshine A. Tayaotao, RPh 2010 5 of 14

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These are nucleoproteins; they are combinations of nucleic acid (DNA) and various proteins. 15% by mass DNA and 85% by mass proteins

FIGURE 5A Chromosomes

DNA REPLICATION Process by which DNA molecules produce exact duplicates of themselves. Semi-conservative replication o o Produces two new DNAs (daughter strand DNAs) identical to each other and exact copies of the original parent DNA Strands are identical to original due to complementary base pairing Complementary base pairing ensures the correct placement of bases in the new DNA strands

Helicase o Unwinds/uncoils DNA helix and splits double strand

Single-Strand Binding Proteins (SSB) o A single-strand binding-protein stabilizes the separated strands, and prevents them from recombining, so that the polymerization chemistry can function on the individual strands.

DNA polymerase o o o Energy is provided to join each new nucleotide to the backbone of a growing DNA strand Catalyzes the replication process at each of these open DNA sections (replication forks) Catalyzes only phosphodiester bonds between the 5-phosphate of one of the nucleotide and the 3-hydroxyl of the next, which means that DNA polymerases have to move in opposite directions along the separated strands of DNA Binds complementary bases a. Leading strand (5 to 3 direction) Replication is continuous

b. Lagging strand (3 to 5 direction) Synthesized in the opposite direction Formed in short segment of 100-200 nucleotides (Okazaki fragments) Page

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Grows in direction of 53 because DNA polymerase III only works in the 53

RNA Primase o Places an RNA primer near fork o Bonds the RNA nucleotides together RNA primer o Short sequence of RNA nucleotides, complementary to a small, initial section of the DNA strand being prepared for replication o Adds short RNA strand to DNA o o Made by adding complimentary RNA nucleotides to the lagging DNA strand by hydrogen bonding of the bases Enzymatically removed and replaced with an appropriate sequence of DNA nucleotides

FIGURE 5A Initiation of Replication by RNA primer and RNA primase

DNA polymerase III o Initiates replication process; Bonds DNA nucleotides to the RNA primer o Adds nucleotides in 5 3 direction (lagging strand)

FIGURE 5B Formation of Okazaki fragments

DNA polymerase I o Removes the RNA primers creating Okazaki fragments and replaces it with DNA Okazaki fragments DNA ligase o Adds DNA nucleotides to fill the gaps between Okazaki fragments and connect lagging strand fragments by creating sugar-phosphate bond

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FIGURE 5C Replication fork, Leading and Lagging strand

FIGURE 5D Summary of DNA Replication

RIBONUCLEIC ACIDS (RNA) a. RIBOSOMAL RNA (rRNA) o Combines with a series of proteins to form complex structures called ribosomes, that serves as the physical sites or platform for protein synthesis b. MESSENGER RNA (mRNA) o Carries genetic information (instructions for protein synthesis) from DNA to ribosome o Primary Transcript RNA (ptRNA) Material from which messenger RNA (mRNA) is made c. TRANSFER RNA (tRNA) o Delivers specific individual amino acids to the ribosomes, the sites of protein synthesis c. OVERVIEW OF PROTEIN SYNTHESIS

FIGURE 6A Protein Synthesis

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1. TRANSCRIPTION Process by which DNA directs the synthesis of RNA molecules that carry the coded information needed for protein synthesis Begins when the section of a DNA molecule that contains the gene to be copied unwinds, which contains a transcription bubble Steps: a. A portion of the DNA double helix unwinds, exposing some bases. The unwinding process is governed by the enzyme RNA polymerase rather than by DNA helicase (replication enzyme) b. Free ribonucleotides align along one of the exposed strands of DNA bases forming new base pairs. In this process, U rather than T aligns with A in the base-pairing process. Because ribonucleotides rather than deoxyribonucleotides are involved in the base-pairing, ribose rather than deoxyribose, becomes incorporated into the new nucleic acid backbone. c. RNA polymerase links the aligned ribonucleic acids. d. Transcription ends when the RNA polymerase encounters a sequence of bases that is read as a stop signal. The newly formed RNA molecules and the RNA polymerase enzyme are released, and the RNA polymerase enzyme are released and the DNA then rewinds to reform the original double bonds. In DNA-RNA base pairing, the complementary base pairs are: o DNA o RNA RNA molecules contain the base U instead of T o The primary function of mRNA molecules is to direct the synthesis of the many different proteins needed for cellular function. Within a DNA strand are instructions for the synthesis of numerous mRNA molecules. During transcription, the DNA molecule unwinding is controlled by RNA polymerase and occurs only at the particular spot where the appropriate base sequence is found for the mRNA (and protein) of concern. Such short segments of DNA, containing instructions for the formation of particular mRNAs are called genes o Gene is a segment of DNA molecule that contains the base sequence for the production of single specific protein molecule. It is now known that not all bases in a gene convey genetic information. Instead, a gene is segmented in portions called exons that contain genetic information and portions called introns that convey genetic information o EXON DNA segment that conveys (codes for) genetic information and helps express a genetic message. o INTRON DNA segment that does not convey (code for) genetic information and interrupt a genetic message. Both exons and introns of a gene are transcribed during production of primary transcript RNA (ptRNA) or pre-mRNA. The ptRNA is then edited, under the direction of enzymes to remove introns. The remaining exons are joined together to form a shortened RNA strand that carries the genetic information of the transcribed gene. This edited RNA is the messenger RNA (mRNA) that serves as a blueprint for protein assembly. 2. TRANSLATION Process by which the codes within the RNA molecules are decipheral and a particular molecule is formed. Process whereby the nucleotide sequence in an mRNA molecule specifies the amino acid sequence of protein. Ribosome in the cytoplasm carries out translation. CODON ANTICODON a. ACTIVATION OF tRNA o An amino acid interacts with an activator molecule (ATP) to form a highly energetic complex. This complex that reacts with an appropriate tRNA molecule to produce an activated tRNA molecule that has an amino acid covalently bonded to it at its 3 end through an ester linkage b. INITIATION o Begins with mRNA which attaches itself to the surface of a small ribosomal subunit such that its first codon, which is always the initiating codon AUG; occupies a site called the P site (peptidyl site) c. ELONGATION o The polypeptide continues to grow by way of translation until all necessary amino acids are in place and bonded to each other. d. TERMINATION o Appearance in the mRNA codon sequence of one of the three stop codons (UAA, UAG, or UGA) terminates the process e. POST-TRANSLATION PROCESSING Prepared by: Sunshine A. Tayaotao, RPh 2010 9 of 14 Page

Some modification of proteins usually occurs after translation

THE GENETIC CODE 1. The genetic code is highly degenerated that is many amino acids designated by more than one codon. Codons that specify the same amino acids are called synonyms 2. There is a pattern to the arrangement of the genetic code table 3. The genetic code is almost universal, the same codon specifies the same amino acid whether the cell is a bacterial cell, a corn plant cell, or a human cell 4. An initiation codon exists o Stop codons o Start codon First Position (5 end) A A Lys Asn Lys Asn Gln His Gln His Glu Asp Glu Asp Stop Typ Stop tyr Second Position C G U Thr Arg Ile Thr Ser Ile Thr Arg Met/Start Thr Ser Ile Pro Arg Leu Pro Arg Leu Pro Arg Leu Pro Arg Leu Ala Gly Val Ala Gly Val Ala Gly Val Ala Gly Val Ser Stop Leu Ser Cys Phe Ser Trp Leu Ser Cys Phe Third Position (3 end) A C G U A C G U A C G U A C G U

TABLE 3 mRNA codons: The Genetic Code for Amino Acids

mRNA 5- GCC AUG GUA AAA UGC GAC CCA 3

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FIGURE 6B Steps in Transcription

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FIGURE 6B Steps in Transcription

GENETIC MUTATIONS: Occurs if one of the code letters is omitted or if one is added or if the order of the code letters is rearranged. Such a change in the sequence of nucleotides may: 1. Give no detectable effect, particularly of the change is in the third letter of the code. 2. Cause a different amino acid to be incorporated into the chain (the results may be acceptable, or totally unacceptable to the function of the protein) 3. Cause the termination of the chain prematurely so that the protein cannot normal. Causes: 1. Exposure to radiation 2. Because of naturally occurring radiation and cosmic rays 3. Exposure to certain chemicals Types: 1. Frameshift Mutation Prepared by: Sunshine A. Tayaotao, RPh 2010 12 of 14 Page

Usually severe, producing a completely nonfunctional protein.

2. Point Mutation Involves a single nucleotide, thus a single amino acid. 3. Silent, Missense, and Nonsense Mutations a. Missense mutation b. Nonsense mutation c. Silent mutation GENETIC DISEASES: Mostly caused by defective gene resulting in a loss of activity of some enzyme 1. CYSTIC FIBROSIS Thick mucus develops and results in bronchia. Obstruction early in childhood infection occurs and becomes difficult to eradicate even with antibiotics. When such an infection develops in the lungs, the subsequent inflammatory reaction results in the destruction of bacteria. Leukocytes and tissue, the process of cellular destruction releases DNA, which in turn substantially increases the viscosity of the mucus. Treatment DNAse, an enzyme that degrades extracellular DNA nut has no effect on the DNA within intact cells 2. PHENYLKETANURIA (PKU) Results when the enzyme phenylalanine hydroxylase is absent. A person with PKU cannot convert phenylalanine to tyrosine, and so the phenylalanine accumulates in the body, resulting in injury to the nervous system. In children up to 6, an accumulation of phenylalanine leads to retarded mental development. It can be readily diagnosed from a sample of blood or urine. Treatment Consists of giving the affected person a diet low in phenylalanine and adding tyrosine to the diet. 3. SICKLE CELL ANEMIA Sickle cells (containing hemoglobin) are fragile than normal red blood cells which leads to anemia. They can also occlude capillaries, leading to thrombosis. Sickle cells in the capillaries cause slowing and sludging of the red blood cells in the capillaries, with resulting hypoxia of the tissues which in turn produces symptoms like fever, swelling, and pain in various parts of the body. A defective Hgb from a mutation in a gene in chromosome 11 decreases the oxygen carrying ability of RBC which takes on a sickle shape, causing anemia and plugged capillaries from RBC segregation. 4. GALACTOSEMIA Results from the lack of enzyme uridyl transferase, which catalyzes the formation of glucose from galactose. This disease may result in an increased concentration of galactose in the blood. Galactose in the blood is reduced in the eye to galacticol, which accumulates and causes a cataract, even liver failure and mental retardation will occur. Transfer of enzyme requuired for the metabolism of glucose-1-phosphate is absent. Accumulation of Gal-1-P leads to cataracts and mental retardation. Treatment Administration of galactose free diet 5. WILSONS DISEASE Caused by the bodys failure to eliminate excess Cu 2+ ions because of a lack of ceruplasmin or failure in the bonding of copper ions to the copper-bonding globulin or both. In this disease, copper accumulates in the liver, kidneys and brain. Increased copper in the kidneys may lead to damage of the renal tubules, leading to increased urinary output of amino acids and peptides. 6. ALBINISM Caused by lack of enzyme tyrosinase, which is necessary for the formation of melanin, the pigment of the hair, skin, and eyes. Although the disease is not serious, persons affected by it are very sensitive to sunburn. 7. HAEMOPHILIA Caused by a missing protein, an antihemophilic globulin, which is important in the normal clotting process of the blood. Consequently, any cut may be life threatening to hemophiliacs, but the primary damage is the crippling effect of repeated episodes of internal bleeding into body joints. One or more defective blood clotting factors lead to poor coagulation, excessive bleeding, and internal hemmorhages. Prepared by: Sunshine A. Tayaotao, RPh 2010 13 of 14 Page

8. MUSCULAR DYSTROPHY (MD) Duchennes muscular dystrophy, one of 10 forms of MD, is caused by the lack of protein called dystrophin, as caused by a mutation in the X chromosome. This disease primarily affects boys at about age 5 with death by age 20. It causes progressive weakness and muscle wasting. 9. DOWN SYNDROME The leading cause of mental retardation, occuring in about 1 in every 100 live births, although the mothers age strongly influences its occurrence. Mental and physical problems including heart and eye defects are the result of the formation of three chromosomes, usually Chromosome 21, instead of a pair. 10. FAMILIAL HYPOCHOLESTEREMIA A mutation of a gene on chromosome 19 results in high cholesterol levels that leads to early Coronary Heart Disease in people 30-40 years old. 11. HUNTINGTONS DISEASE (HD) Appearing in middle age, affects the nervous system, leading to total physical impairment. It is the result of a mutation in a gene on chromosome 4, which can now be mapped to test people in families with HD. 12. TAY-SACHS DISEASE Hexosamindase A is defective, causing an accumulation of gangliosides resulting in mental retardation, loss of motor control, and early death. 1. 2. 3. 4. TREATMENT OF GENETIC DISEASES: Correct the metabolic consequences of the disease by supplying the missing product. Replace the missing enzyme or hormone. Remove excessive stored substances Correct the major genetic abnormality.

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