Nucleic Acids and Nucleosides

• Subject of nucleic acids was first investigated in 1869
• Friedrich Miescher, a 25-year-old Swiss chemist, isolated nuclei from pus cells in 1869
• The nuclei contained an unknown phosphate-rich substance, which Miescher named nuclein
• Nuclein differed from carbohydrates, proteins, and fats
• Nuclein exhibited acidic properties

Nucleic Acids

• Altmann renamed nuclein to nucleic acid in 1899
• Nucleic acids are named for their initial discovery within the nucleus and the presence of phosphate groups
• Nucleic acids determine heredity in living organisms
• They are macromolecules in living cells, either free or bound to proteins (nucleoproteins)
• Nucleic acids are biopolymers of high molecular weight, with mononucleotides as repeating units
• They contain carbon, hydrogen, oxygen, nitrogen, and phosphorus
• Nucleic acids are amongst the most important biomolecules
• They are found in all living things and are collectively known as DNA and RNA

Nucleosides

• Nucleosides result from linking a sugar with a purine or pyrimidine base through an N-glycosidic linkage
• Purines bond to the C1 carbon of the sugar at their N9 atoms
• Pyrimidines bond to the C1 carbon of the sugar at their N1 atoms

Bases, Sugars and Nucleosides

Phosphoric Acid

• Phosphoric acid can exist as mono-, di-, or triphosphates
• Phosphates bond to the c3 or c5 atoms of the sugar

Nitrogenous Bases

• Nitrogenous bases include pyrimidines and purines

Pyrimidines

• Pyrimidines include single heterocyclic ring structures
• Cytosine exists in both RNA and DNA
• Thymine exists only in DNA
• Uracil exists only in RNA
• Nucleotide base carbons are denoted as 1,2, etc.
• Pyrimidine bases exist in lactam and lactim forms

Lactam and Lactim Forms

• The group -HN-CO- forms a lactam (keto)
• Isomerization to -N=C-OH forms a lactim (enol)
• At physiological pH, the lactam form predominates
• Cytosine is chemically 2-deoxy-4-amino pyrimidine
• Cytosine exists in lactam or lactim forms
• Cytosine is found in all NAs except in the DNA of certain viruses
• Uracil is chemically 2,4-dideoxy pyrimidine
• Uracil is confined to RNA only
• Thymine (5-methyl-Uracil) is 2,4-dideoxy-5-methyl pyrimidine
• Thymine is found only in DNA
• Small amounts of thymine have been found in t-RNA

Purines

• Purines have heterocyclic double-ring structures
• Purines are fusion products of a pyrimidine ring with imidazole ring
• Adenine and Guanine are both in DNA and RNA

Adenine and Guanine

• Adenine is chemically 6-aminopurine
• Guanine is 2-amino-6-oxypurine
• Guanine can exist as lactam and lactim forms by isomerization

Minor Bases

• Minor bases are unusual and may exist with no known function
• Some are found in the NAs of bacteria and viruses
• Many are found in DNA and tRNAs of both pro- and eukaryotes

Examples of Modified Bases

• Bacteriophages contain 5-hydroxy methyl cytosine
• mRNA of mammalian cells contain N6-methyl adenine and N7-methyl guanine, and N6-N6-dimethyl adenine
• In plants, purine derivatives include theophylline (1,3-dimethyl xanthine in tea), theobromine (3,7-dimethyl xanthine in cocoa), and caffeine (1,3,7-trimethyl xanthine in coffee)

Pentose Sugar

• Numbering of carbon on a pentose sugar is "primed"

Nucleotides

• Nucleotides result from linking one or more phosphates with a nucleoside onto the 5' end of the molecule through esterification
• The purine nucleosides end in “-sine” like Adenosine and Guanosine
• The pyrimidine nucleosides end in "-dine" Thymidine, Cytidine, Uridine

Nucleotide Nomenclature

• Use the nucleoside name from above and add "mono-", "di-", or "triphosphate"

Importance of Nucleotides & Nucleosides

• Apart from DNA & RNA, they have diverse functions
• Adenosine nucleotides include ATP, ADP, AMP, and cAMP
• ATP is the storage battery of tissues
• ATP is the energy currency of the cell
• ATP is a storehouse of energy
• 2~P releases energy (7.6Kcal), resulting in endergonic reactions
• Used in gluconeogenesis, FA, Creatine, and SAM synthesis
• Used for phosphotransferases
• Used to synthesize 'active' sulfate (PAPS)
• Converted to ADP, AMP, and 3',5'-cAMP

Adenosinediphosphate (ADP)

• It is a 1º Pi acceptor in oxidative phosphorylation(OP)→ADP+Pi=ATP
• It controls cellular respiration, muscle contraction, etc
• It is an activator of the enzyme Glutamate dehydrogenase
• serves as the sulfate donor for proteoglycans and conjugations of drugs
• S-adenosylmethionine is a methyl group donor
• Serves as an allosteric regulator and an energy source for protein synthesis

Second Messengers

• cAMP and cGMP act as the second messengers
• cAMP is a secondary messenger for many hormones
• cGMP serves as a 2nd messenger in response to nitric oxide, particularly during the relaxation of smooth muscles

UDP-Sugar Derivatives

• Participates in: Sugar epimerization
• Participates in: Biosynthesis of glycogen & glucosyl disaccharides, oligosaccharides of glycoproteins & proteoglycans

UDP-Glucuronic Acid Forms

• Is the urinary glucuronide of bilirubin
• Conjugates many drugs such as aspirin

CTP

• CTP participates in the biosynthesis of phosphoglycerides, sphingomyelin, & other substituted sphingosines

Synthetic Nucleotide Analogs and Chemotherapy

• Synthetic analogs of purines, pyrimidines, nucleosides, and nucleotides are made by altering the heterocyclic ring or the sugar moiety
• Has numerous applications in clinical medicine
• Their toxic effects reflect either inhibition of enzymes essential for NA synthesis or their incorporation into nucleic acids
• This results in disruption of base-pairing

Common Agents

• Allopurinol treats hyperuricemia & gout, inhibits purine biosynthesis & xanthine oxidase activity
• Cytarabine is used in chemotherapy of cancer
• Azathioprine is catabolized to 6-mercaptopurine
• Azathioprine is employed during organ transplantation to suppress immunologic rejection

Function of Nucleic Acids

• Nucleic acids serve as the genetic material of living organisms
• They are involved in the storage, transfer, and expression of genetic information
• Nucleic acids contain all the information needed for the formation of individuals or organisms
• They determine the physical fitness of an individual to life
• Some act as enzymes and coenzymes

Polynucleotides

• Nucleic acids are polynucleotides (nucleic acid)
• Polynucleotides are directional macromolecules
• Phosphodiester bonds link the 3'- and 5'-C's of adjacent monomers
• Each end of a nucleotide polymer thus is distinct
• The "5'- end" and the "3'- end" of polynucleotides have different properties
• The 5'- end is the one with a free/phosphorylated 5'-OH

Polynucleotide Primary Structure

• The base sequence/primary structure of a polynucleotide can be represented as shown below
• The phosphodiester bond is represented by P or p
• Bases are represented by a single letter
• Pentoses are represented with a vertical line
• All phosphodiester bonds are 5'→3'
• A more compact notation is possible
• An example compact notation is pGpGpApTpCpA

Compact Representation

• Indicates that the 5'-OH is phosphorylated, but not the 3'-OH
• The most compact representation shows only the base sequence
• With the 5'- end on the left and the 3'- end on the right
• The phosphoryl groups are assumed but not shown
• GGATCA is an example

Dinucleotides

• The 5'-phosphoryl group of a mononucleotide esterifies the 3'-OH of the pentose of a 2nd nucleotide
• Forming a phosphodiester
• This forms a dinucleotide in which the pentose moieties are linked by a 3'→5' phosphodiester bond
• Forming an RNA & DNA backbone
• Dinucleotide formation includes elimination of water between two mononucleotides

Nucleic Acids

• Nucleic Acids (NAs) are either DNA or RNA
• Double-stranded, helical polymers of deoxyribonucleotides found in the nucleus
• Contains Storage of the genetic material
• Single stranded, non-helical polymers of ribonucleotides found in the nucleus and cytoplasm
• Transfers the genetic information from nucleus to cytoplasm for protein synthesis

RNAs

• Messenger RNA
• Is a copy of selected regions of the DNA
• It carries the genetic message from the nucleus to the cytoplasm
• m-RNA acts as the template for protein synthesis.
• Ribosomal RNA
• rRNA, along with proteins, forms the ribosome
• Ribosomes are the site of protein synthesis
• Some ribosomes have catalytic and coenzyme functions
• Transfer RNA
• tRNA transfers the amino acids from the cytoplasm to the site of protein synthesis

DNA Primary Structure:

• It is the linear sequence of building nucleotide units

DNA Backbone

• Phosphate and sugar form the backbone of the DNA molecule
• The bases form the "rungs"
• There are four types of nitrogenous bases
• They bounds with its specific base in two poly peptide run in opposite direction

Base Pairing

• Each base will only bond with one other specific base
• Due to this complementary base pairing, the order of the bases in one strand determines the order of the bases in the other strand
• If A is on one strand, T is on the other
• If C is on one strand, G is on the other
• The base pairs are perpendicular to the axis of the helix
• One polynucleotide chain of the DNA double helix is always the complement of the other

Chargaff's Rule

• The specific base pairing in DNA leads to Chargaff's Rule
  • In double stranded DNA, amount of A = amount of T and the amount of G = amount of C
  • The total amount of purines equals the total amount of pyrimidines

Erwin Chargaff

• Erwin Chargaff's findings by 1950 suggested base-pairings of A-T & G-C
• He shared his findings with Watson and Crick in 1952
• "Chargaff's rule" is A = T & C = G

Double Helix

• The base pairs are held together by hydrogen bonds
  • Two between A and T
  • Three between G and C
• Hydrogen bonds stabilize its structure
• In addition hydrophobic interactions between the stacked bases are important

Watson and Crick

• James Watson and Francis Crick worked together at Cavendish Laboratory in Cambridge
• Worked to determine DNA structure
• They used work from Franklin, Wilkins, and Chargaff to determine the double helix shape
• The two chains are coiled around a common axis, called the axis of symmetry
• The chains are antiparallel
• The hydrophilic deoxyribose-phosphate backbone of each chain is on the outside of the molecule, whereas the hydrophobic bases are stacked inside
• The overall structure resembles a twisted ladder
• Spatial relationships between the two strands in the helix creates major and minor grooves
• Grooves provide access for the binding of regulatory proteins

Eukaryotic DNA Organization

• Eukaryotic DNA is associated with tightly bound basic proteins, called histones
• These serve to order the DNA into fundamental structural units, called nucleosomes
• Nucleosomes are further arranged into more complex structures that condense the DNA molecules into chromosomes
• Chromosomes can be segregated during cell division
• The complex of DNA and protein found inside eukaryotic cells is called chromatin

Histones

• There are five classes of histones: H1, H2A, H2B, H3and H4
• These small proteins are positively charged at physiologic pH due to their high content of lysine and arginine
• Due to their positive charge, they form ionic bonds with negatively charged DNA
• Histones, along with positively charged ions such as Mg2, help neutralize the negatively charged DNA phosphate groups

RNA Types

• RNA is termed Ribonucleic acid
• Its bases consist of Cytosine, guanine, adenine and uracil
• It is typically Single stranded
• rRNA (ribosomal RNA) makes up about 60% of ribosomal structure
• mRNA (messenger RNA) record information from DNA and carry it to ribosomes
• tRNA (transfer RNA) delivers amino acids to proteins at the ribosome to extend the chains

RNA Structure

• RNA Primary structure is the sequence of ribonucleotides in chain

• Secondary structure

  • Even though most RNAs are single stranded, they exhibit extensive secondary structures, including intramolecular double-stranded regions

• These secondary structures are important to their function

• One of the most common types is a hairpin loop

• Hairpin loops are produced by intramolecular base pairing

• They occur between complementary nucleotides within a single RNA molecule

• Tertiary structure involves the folding of the molecule into a 3D shape

• This is stabilized by hydrophobic bonds and H-bonds, producing a compactly coiled globular structure

Transfer RNA

• Are Adapter molecules
• Act as adapters for the translation of the sequence of nucleotides of the m RNA in to specific amino acids
• There are at least 20 species of t RNA
• Each corresponds to each of the 20 amino acids required for protein synthesis
• The nucleotide sequence of all the t RNA molecules allows extensive complimentarity
• Generates a secondary structure.

tRNA Secondary Structure

• Each single t-RNA shows extensive internal base pairing and acquires a clover leaf like structure
• The structure is stabilized by hydrogen bonding between the bases and is a consistent feature
• All t-RNA contain 5 main arms or loops
  • a) Acceptor arm
  • b) Anticodon arm
  • c) DHU arm
  • d) T C arm
  • e) Extra arm

Structural features

• Bases that commonly occur in a position are indicated by letters
• The locations of modified bases are indicated, such as:
  • Dihydrouridine (D)
  • Ribothymidine (T)
  • Pseudouridine (ψ)

Acceptor Arm

• Is at the 3 end
• It has 7 base pairs
• The end sequence is unpaired Cytosine,Cytosine-Adenine at the 3 end
• The 3 OH group terminal of Adenine binds with carboxyl group of amino acids
• The tRNA bound with amino acid is called Amino acyl t RNA
• CCA attachment is done post transcriptionally
• The carboxyl group of amino acid is attached to 3OH group of Adenine nucleotide of the acceptor arm

Anticodon Arm

• Lies at the opposite end of the acceptor arm
• 5 base pairs long
• Recognizes the triplet codon present in the m RNA
• Base sequence of anticodon arm is complementary to the sequence of the m RNA codon
• Due to complimentarity it can bind specifically with m RNA by hydrogen bonds

DHU Arm

• It has 3-4 base pairs
• Serves as the recognition site for the enzyme (amino acyl t RNA synthetase) that adds the amino acid to the acceptor arm
• ΨC arm
  • This arm is opposite to DHU arm
  • It contains pseudo uridine, that's why it is so named
  • It is involved in the binding of t RNA to the ribosomes

Variable Arm

• The Extra arm or Variable arm
• Generally makes up about 75 % of t RNA molecules possess
• For about 3-5 base pairs are present the t-RNA is class 1
  • Majority t -RNA belong to class 1.
• The t –RNA belonging to class 2 have long extra arm; 13-21 base pairs in length
• The L shaped tertiary structure is formed by further folding of the clover leaf due to hydrogen bonds between T and D arms
• The base paired double helical stems are arranged into two double helical columns, continuous and perpendicular to one another

DNA vs RNA