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Chemistry of nucleic acids

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Chemistry of nucleic acids

  1. 1. R.C. Gupta Professor and Head Department of Biochemistry National Institute of Medical Sciences Jaipur, India Chemistry of Nucleic Acids (DNA and RNA)
  2. 2. Nucleic acids are known as information molecules They store genetic information The nucleic acids are of two types: • Deoxyribonucleic acid (DNA) • Ribonucleic acid (RNA)
  3. 3. In most of the organisms, genetic information is present in DNA In some viruses, it is present in RNA
  4. 4. The information present in DNA concerns the synthesis of proteins This information is present in a coded form Different types of RNA are required to synthesize the proteins
  5. 5. The nucleic acids are large polymers Their building blocks are nucleotides DNA is a polymer of deoxyribonucleotides RNA is a polymer of ribonucleotides
  6. 6. DNA is present in the nucleus (in eukaryotes, some DNA is present in the mitochondria also) RNA is mainly extranuclear
  7. 7. Deoxyribonucleic acid (DNA) Miescher named it as nuclein as it was present in the nucleus The earliest evidence about the presence of DNA in cells was obtained by Friedrich Miescher in 1869
  8. 8. Richard Altmann found that nuclein was an acid; hence he named it as nucleic acid It 1919, Phoebus Levene identified the components of nucleic acid
  9. 9. The components of nucleic acid were found to be bases, sugars and phosphate The nucleic acid containing deoxyribose was named DNA The nucleic acid containing ribose was named RNA
  10. 10. The role of DNA as genetic material was first shown by Griffith in 1928 He conducted his experiments on pneumococci Pneumococci are of two types Encapsulated Non-encapsulated EMB-RCG
  11. 11. Encapsulated pneumococci form smooth colonies, and are pathogenic Non-encapsulated pneumococci form rough colonies, and are non-pathogenic Each type produces its own kind of offspring
  12. 12. Griffith transferred nuclear material from encapsulated pneumococci into non- encapsulated pneumococci When the non-encapsulated pneumococci divided, the daughter cells had capsules, and were pathogenic Thus, it was proved that genetic information is present in the nucleus
  13. 13. Griffith termed nuclear material as the transforming principle However, nuclear material contains a variety of compounds Due to their diversity, proteins and nucleic acids became the prime candidates for ‘transforming principle’
  14. 14. Avery et al (1944) treated nuclear material with enzymes that hydrolyse DNA or RNA or proteins Hydrolysis of DNA destroyed the trans- forming activity of nuclear matter but hydrolysis of RNA or proteins did not
  15. 15. Avery et al concluded that genetic information is present in DNA Some researchers still speculated that genetic information might be present in nuclear proteins and not in DNA This doubt was cleared by Hershey and Chase in 1952
  16. 16. Martha Chase Alfred Hershey
  17. 17. Hershey and Chase conducted their studies on T2 bacteriophage, a DNA virus T2 bacteriophage is made up of a DNA core surrounded by a protein coat T2 bacteriophage infects the bacterium, E. coli
  18. 18. The bacteriophage multiplies inside the infected E. coli When the number of viruses becomes too large, the bacterial cell ruptures The viruses are released
  19. 19. The experiment was repeated using 35S as label This time, viral proteins were labelled and the virus was allowed to infect E. coli
  20. 20. This showed that when the virus infected the bacterium, only the viral DNA entered the bacterial cell and not the proteins Since progeny viruses were formed and proteins surrounded DNA, viral DNA must have directed the synthesis of new proteins This established the role of DNA as the genetic material
  21. 21. Structure of DNA Chargaff (1950) studied the base composition of DNA obtained from diverse sources He found that number of A residues was equal to T residues and number of C residues was equal to G residues in every DNA A = T C = G
  22. 22. Wilkins and Franklin did extensive x-ray crystallographic studies on DNA Maurice Wilkins Rosalind Franklin They showed that DNA has a helical structure
  23. 23. X-ray crystallographic images of DNA Typical of helical structure Typical of helical structure
  24. 24. Watson Crick James Watson and Francis Crick analyzed: X-ray crystallographic data And other facts Structures of purine and pyrimidine bases Chargaff’s observations
  26. 26. Size of purines and pyrimidines
  27. 27. Watson & Crick proposed a model of DNA structure
  28. 28. Watson & Crick announced their discovery on Feb 28, 1953 (published in April, 1953) Their model was consistent with all the known features of DNA
  29. 29. Each strand of DNA is a polymer of mononucleotides The successive nucleotides in a strand are linked by 3', 5'-phosphodiester bonds According to Watson & Crick model, DNA is a double-stranded helix
  30. 30. The bases present in DNA are: Adenine Guanine Cytosine Thymine
  31. 31. Each strand has got a polarity or direction Each strand has a 3'-end and a 5'-end S − P − S − P − S − P − S − P B B B B B B B B P − S − P − S − P − S − P − S 5’ End 5’ End 3’ End 3’ End
  32. 32. The two strands are anti-parallel i.e. they are parallel but run in opposite directions At 5'-end, –OH group attached to carbon atom 5 of the sugar is not esterified At 3'-end the –OH group attached to carbon atom 3 of the sugar is not esterified
  33. 33. There are two hydrogen bonds between adenine and thymine (A=T), and three between guanine and cytosine (G≡C) All the bases in the molecule take part in hydrogen bonding with complementary bases on the opposite strand The bases are present in the interior of the molecule while the sugar and phosphate groups are present on the outer side
  34. 34. 5’ 3’ 3’ 5’ (b)(a)
  35. 35. The two strands are not straight They are wound around each other to form a double helix
  36. 36. The double helix looks like a twisted ladder
  37. 37. Each turn of the helix contains ten base pairs It has a pitch of 3.4 nm The diameter of the helix is 2 nm The helix is right-handed
  38. 38. Two grooves are seen in the double helix These are termed as the major groove and the minor groove The grooves are present between the glycosidic bonds on the opposite strands
  39. 39. 3.4 nm (pitch) 3.4 nm (Pitch) 2 nm (Diameter)
  40. 40. Richard Dickerson found a slightly different structure while studying DNA crystals The structure found by Dickerson was named as A-DNA, and that described by Watson and Crick was termed as B-DNA A third type of structure was found by Alexander Rich which was named as Z-DNA
  41. 41. B-DNA is the commonest type of DNA A-DNA is formed when the environment is less humid Z-DNA is formed when pyrimidine and purine bases alternate in a DNA strand
  42. 42. B-DNA A-DNA Z-DNA
  43. 43. A-DNA B-DNA Z-DNA EMB-RCG Direction of Right- Right- Left- helix handed handed handed Minor groove Wide Narrow Very narrow Major groove Narrow Wide Flat Glycosidic bond syn anti syn (purines) anti (pyrimidines) Number of base pairs per turn 11 10 12 Rise per base pair 0.25 nm 0.34 nm 0.37 nm Rise per turn (pitch) 2.7 nm 3.4 nm 4.4 nm Diameter 2.3 nm 2.0 nm 1.8 nm Important features of A-, B- and Z-DNA
  44. 44. Sense and anti-sense Genetic information is present on one strand of DNA which is known as the sense strand The other strand has a complementary sequence of bases, and is known as the anti-sense strand
  45. 45. During replication, the two strands separate, and each serves as a template A new strand having a complementary base sequence is synthesized on each template strand Thus, the new DNA becomes an exact replica of the original DNA
  46. 46. The DNA is combined with a nearly equal amount of proteins to form nucleoproteins The predominant proteins are histones which are of five types – H1, H2A, H2B, H3 and H4 The histones are basic proteins rich in lysine and arginine
  47. 47. The positively charged amino acids interact with negatively charged phosphate groups of DNA The basic amino acids are present mainly in the N-terminal and C-terminal regions The inner core of histones contains non-polar amino acids which form a globular structure
  48. 48. Two molecules each of histones H2A, H2B, H3 and H4 form an octamer around which DNA is wrapped in two coils to form a nucleosome Histone octamer DNA Nucleosome
  49. 49. Histone octamer DNA H1Chromatosome A nucleosome associated with histone H1 is called a chromatosome
  50. 50. ― Linker DNA Polynucleosome A series of nucleosomes (“beads on a string”) form a polynucleosome The DNA between two nucleosomes is known as linker DNA
  51. 51. Nuclesomes (10 nm wide) condense to form 30 nm wide nucleofilaments In this way, the linear DNA becomes highly compact Even higher compactness is achieved by looping of nucleofilaments The loops associate with some scaffold proteins to form a chromosome
  52. 52. Nucleofilament Chromosome
  53. 53. Many non-histone proteins are also associated with DNA in small amounts They have a bearing on various functions like replication and transcription Human beings have 23 pairs of chromosomes Each chromosome consists of a single molecule of double-helical DNA along with several proteins
  54. 54. Most of the DNA present in a cell is supercoiled (superhelical) In supercoiled DNA, the axis of the double helix is bent and twisted upon itself In negatively supercoiled DNA, the twists are right-handed Most of the naturally occurring DNA is negatively supercoiled
  55. 55. Circular DNA (relaxed) Circular DNA (negatively supercoiled) Circular DNA (positively supercoiled)
  56. 56. Ribonucleic acid (RNA) RNA is also a polymer of mononucleotides The bonds between the mononucleotides are similar to those in DNA However, RNA differs from DNA in some ways
  57. 57. EMB-RCG Differences between RNA and DNA DNA RNA Sugar Deoxyribose Ribose Pyrimidine bases Cytosine and thymine Cytosine and uracil Number of strands Two One Chargaff’s rule Followed Not followed
  58. 58. EMB-RCG The RNA strand may be folded upon itself Intra-strand hydrogen bonds may be formed between complementary bases present in the same strand This may give a double-stranded look in certain regions of the molecule
  59. 59. There are three types of RNA: Messenger RNA (mRNA) Transfer RNA (tRNA) Ribosomal RNA (rRNA)
  60. 60. Structure and functions of different types of RNA EMB-RCG Type Structure Function mRNA Single, uncoiled strand Transmits information from DNA and serves as a template for protein synthesis tRNA Single strand folded back on itself Brings amino acids to ribosomes for protein synthesis rRNA Globular rRNA and proteins make up ribosomes
  61. 61. Messenger RNA Messenger RNA carries message from the nucleus to the ribosomes Genetic information is present on the sense strand of DNA in the form of genes A gene is a part of the sense strand having a specific nucleotide sequence
  62. 62. The gene contains coded information for the synthesis of a particular protein Each mRNA molecule is a transcript of the sense strand of a particular gene Its nucleotide sequence is complementary to that of the sense strand of the gene
  63. 63. EMB-RCG At its 5'-end, mRNA possesses a 7- methylguanosine triphosphate cap The cap helps the protein-synthesizing machinery to identify the mRNA At its 3'-end, it has a poly-A tail made up of several adenylate residues The tail stabilizes the structure of mRNA
  64. 64. The mRNA molecule is initially synthesized in eukaryotes as a precursor The precursor is heterogeneous nuclear RNA (hnRNA) or pre-mRNA hnRNA is much bigger than the final mRNA
  65. 65. hnRNA is processed to form mRNA The cap and tail are added during processing Many nucleotides are removed during processing
  66. 66. Transfer RNA tRNA is made up of about 75 nucleotides Its molecular weight is about 25,000 It transports amino acids from cytosol to ribosomes for protein synthesis
  67. 67. A given tRNA is specific for one amino acid Proteins are synthesized from 20 amino acids Therefore, there are at least 20 species of tRNA
  68. 68. Like other RNAs, tRNA is single-stranded Hydrogen bonds are formed between complementary bases on the same strand This gives rise to secondary and tertiary structures
  69. 69. 5’-End 3’-End Secondary structure is folded upon itself to form an L-shaped tertiary structure
  70. 70. At the 3'-end, the last three nucleotides are –C–C–A The amino acid is attached to the terminal adenylate residue The 3'-end is known as the acceptor arm of tRNA
  71. 71. Pseudouridine (y) loop The tRNA molecule has three loops (or arms) known as: Anticodon loop Dihydrouracil (DHU) loop
  72. 72. The anticodon loop contains a triplet of nucleotides known as anticodon The anticodon is complementary to a codon on the mRNA As there is only one anticodon on a tRNA, it is specific for one amino acid
  73. 73. Ribosomal RNA rRNA is a structural constituent of ribosomes The ribosomes are made up of 40S subunit and 60S subunit in eukaryotes Each subunit is made up of some poly- peptides and some molecules of rRNA
  74. 74. Eukaryotes have four types of rRNA differing in size: 5S rRNA 5.8S rRNA 18S rRNA 28S rRNA
  75. 75. 5.8S, 18S and 28S rRNA are formed from a single 45S precursor The 5S rRNA is formed as such
  76. 76. rRNA molecules are extensively folded
  77. 77. rRNA molecules combine with the poly- peptides to form globular ribosomal subunits At the time of protein synthesis, the two subunits combine to form the ribosome mRNA and tRNAs bind to the ribosome to synthesize a protein EMB-RCG
  78. 78. Ribosome 60S 40S