Lecture 5 Nucleic Acid BIOL 158 PDF

NUCLEIC ACIDS  The central dogma of biosciences DNA RNA Protein Genetic Transmitter of genetic Biocatalyst Information information Molecular machine  Nucleotides and Nucleic acids NUCLEIC ACIDS Deoxyribonucleic acid (DNA) Ribonuclei...

NUCLEIC ACIDS  The central dogma of biosciences DNA RNA Protein Genetic Transmitter of genetic Biocatalyst Information information Molecular machine  Nucleotides and Nucleic acids NUCLEIC ACIDS Deoxyribonucleic acid (DNA) Ribonucleic acid (RNA) Linear polymers of nucleotides NUCLEOTIDES Ribose/Deoxyribose sugar Heterocyclic nitrogenous bases (pyrimidines & purines) Phosphate group  Things to learn from the lecture What are the structures of the nucleotides? How are nucleotides joined together to form nucleic acids? How is information stored in nucleic acids? What are the biological functions of nucleotides and nucleic acids? What are the structures of the nucleotides? Nitrogenous bases: derivatives of pyrimidine or purine Three pyrimidines and two purines are commonly found in cells Purine derivatives Ribose 5 OH HOCH2 O OH HOCH2 O 4 1 2 2 3 OH H OH OH Ribose Deoxyribose β-D-Ribofuranose β-D-2-Deoxyribofuranose Nucleosides (ribose + base) β-N-glycosidic bond Syn and anti conformations of nucleosides NH2 NH2 1N N N N 1 N N N N 9 9 HOCH2 O HOCH2 O OH OH OH OH Syn-conformation syn Anti-conformation anti The anti conformations predominate in nucleic acids, the polymers of nucleotides. Nucleosides are more water soluble than free bases Both kinds of nucleosides are stable in alkali Pyrimidine nucleosides are resistant to acid hydrolysis than purine  O NH N NH O 2 N HN RNA N N 2 HN N NH2 N N N O N O N HOCH2 O HOCH2 O HOCH2 O HOCH2 O OH OH OH OH OH OH OH OH A Adenosine G Guanosine C Cytidine U Uridine NH2 O NH2 O DNA CH3 N HN N N N HN N N NH2 N N O N O N HOCH2 O HOCH2 O HOCH2 HOCH2 O O OH H OH H OH H A OH H G C T Deoxyadenosine Deoxyguanosine Deoxycytidine Deoxythymidine Nucleotides (phosphated nucleosides ) NH2 NH2 Phosphate ester N N N - N N N O O O- P O OCH2 O P OCH2 O O OH H OH OH Adenosine monophosphate Deoxycytidine P AMP monophosphate Phosphoryl group dCMP Structures of common ribonucleotides — AMP, GMP, CMP, and UMP uncommon  Nucleoside diphosphates and triphosphates are nucleotides with two or three phosphate groups Nucleoside 5’-diphosphates/triphosphates (NDPs/NTPs): strong polyprotic acids Form stable complexes with divalent cations: Mg2+, Ca2+ What are the structures and functions of the NA? Nucleic acids are polynucleotides: linear polymers of nucleotides linked by 3’,5’-phosphodiester bridge The base sequence of a nucleic acid is its distinctive characteristic Pi- Sugar-phosphate Side chains: bases backbone DNA: A, G, C, T Two termini of RNA: A, G, C, U Polynucleotides: 5’-Pi (5’- ) 3’- OH (3’- ) Direction of the 5’-end: pTpGpCpAp polynucleotide 3’-end: ApCpGpTp chain -OH 5’-TGCA…… From 5’- to 3’- The different classes of a nucleic acid: DNA and RNA What is the different between structures of DNA and RNA? DNA: quite large only a single molecular (chromosome DNA) in simple life Escherichia coli: 2.9 × 109 D (> million nucleotides) Many chromosomes in eukaryotic cells, in two copies principally; also occurs in mitochondria and in chloroplasts. RNA: small Occurs in multiple copies and various forms in cells As biological functions, they are categorized into 3 major types: messenger RNA, transfer RNA, ribosomal RNA (in all lives), and another kind, small nuclear RNA ( only in eukaryotic cells). mRNA, tRNA, rRNA and snRNA (see table 10.2--p.319) The fundamental structure of DNA is a double helix Chargaff’s rules: in late 1940s, Amounts of the four bases: (p.320) Vary from species to species A = T ; G = C (in each X-ray diffraction studies (R Purine (A+G) = pyrimidine (T+C) Franklin and M Wilkins): Helix with two different loops in outside James Watson and Francis Crick at Cambrige University in 1953: DNA was a complementary double helix; Two strands of DNA are held together by the bonding interactions between unique base pairs. Size of DNA: ~bp; ~kb The genetic information in DNA is encoded in digital form Base paring: interaction between bases—H bonds Replication of DNA: two identical progeny molecules by base paring DNA is in the form of enormously long, threadlike molecules E. coli are partially digested and diluted with water DNA in cells occurs in the form of chromosomes Prokaryotic cell: circular form, bind proteins, chromosomes are no ordered structure Eukaryotic cell: Basic unit--nucleosome Histones, a class of Arg- and Lys-rich basic protein, interact ionically with the anionic phosphate groups of DNA backbone nucleosome Various forms of RNA serve different roles in cells mRNA, tRNA, rRNA, and small nuclear (sn) RNA 1. Messenger RNA carries the sequence information for synthesis of a protein Transcription : synthesis of RNA from DNA by RNA polymerase Prokaryotes Gene expression Translation : synthesis of protein from RNA by ribosome Eukaryotes Heterogeneous heavy nuclear RNA, hnRNA Gene expression 2. Ribosomal RNA provides the structural and functional foundation for ribosomes 3. Transfer RNAs carry amino acids to ribosomes for use in protein synthesis 73~94 residues (bases) 4. Small nuclear RNAs mediate the splicing of Eukaryotic gene transcripts (hnRNA) into mRNA Contain 100~200 nucleotides, found in stable complexes with specific proteins forming small nuclear ribonucleoprotein particles, or snRNPs, are important in the processing of hnRNA. 5. Small RNAs serve a number of roles, including post- transcriptional gene silencing Only 21~28 nucleotides, can target DNA or RNA through complementary base pairing—direct readout. Small interfering RNA (siRNA): disrupt gene expression by binding mRNA complementally to form double-stranded RNA, which is easily degraded and eliminating the mRNA. Are nucleic acids susceptible to hydrolysis? RNA is susceptible to hydrolysis by base, but DNA is susceptible to hydrolysis by acid DNA: HCl—hydrolyzing purine glycosidic bond (apurinic acid) RNA: NaOH—randomly hydrolyzing glycosidic bond The enzymes that hydrolyze nucleic acids are phosphodiesterases  Nucleases differ in their specificity for different forms of nucleic acid DNase, RNase only act on DNA or RNA respectively Restriction enzymes are nucleases that cleave double-stranded DNA molecules There are types I, II and III restriction endonucleases Type I and III require ATP to hydrolyze DNA, Type I cleave DNA randomly, Type III cut DNA in specific sequence. Type II restriction enzymes: no need for ATP, specific recognition sequences are typically 4 or 6 nucleotides in length and have a two fold axis of symmetry  Type II restriction endonuclease EcoRI 5’—N–N–N–N–N–N–G A–A–T–T–C–N–N–N–N–N–N—3’ : : : : : : : : : : : : : : : : : : 3’—N–N–N–N–N–N–C–T–T–A–A G–N–N–N–N–N–N—5’ “sticky” ends—cohesive ends Restriction endonucleases can be used to map the structure of a DNA fragment  Peptide nucleic acids (PNAs) are synthetic mimics of DNA and RNA The sugar phosphate backbone is replaced by a peptide backbone Stable probe PNAs are resistant to nucleases and also are poor substrates for protease Key points: Structures of purine and pyrimidine bases, nucleosides and nucleotides. Structure of polynucleotide strand. Difference between DNA and RNA (structure and function).