Nucleic Acid Biochemistry PDF

NUCLEIC ACID BIOCHEMISTRY BY: MARY ABIGAIL PAULAN, MSMLS, RMT BSML-3225 PRELIM   DNA  DNA STRUCTURE  DNA REPLICATION  ENZYMES THAT METABOLIZE DNA  PHYSICAL PROPERTIES...

NUCLEIC ACID BIOCHEMISTRY BY: MARY ABIGAIL PAULAN, MSMLS, RMT BSML-3225 PRELIM   DNA  DNA STRUCTURE  DNA REPLICATION  ENZYMES THAT METABOLIZE DNA  PHYSICAL PROPERTIES  RNA  TYPES/STRUCTURES OF RNA OUTLINE  RNA TRANSCRIPTION  RNA-METABOLIZING ENZYMES BSML-3225 PRELIM  NONCODING RNAs RESEARCHERS WHO MADE IT POSSIBLE… Friedrich Phoebus Levene Erwin Chargaff Miescher Discovery of Investigates the "Chargaff's “DNA” Structure of DNA rule” BSML-3225 PRELIM JAMES WATSON & FRANCIS CRICK  Proposed the Double Helix structure BSML-3225 PRELIM BSML-3225 PRELIM  “ T HE BEG INNING… BSML-3225 PRELIM  DNA Deoxyribonucleic acid BSML-3225 PRELIM STRUCTURE Nitrogenou s Base Pentose Sugar Phosphate Group BSML-3225 PRELIM NITROGENOUS BASE BSML-3225 PRELIM PENTOSE SUGAR BSML-3225 PRELIM NUCLEOSIDE BSML-3225 PRELIM BSML-3225 PRELIM BSML-3225 PRELIM NUCLEOTIDE BSML-3225 PRELIM BSML-3225 PRELIM BSML-3225 PRELIM THE DOUBLE HELIX FORMATION BSML-3225 PRELIM THE DOUBLE HELIX FORMATION BSML-3225 PRELIM BSML-3225 PRELIM CHARGAFF’S RULES  Two long polynucleotide chains are coiled around a central axis, forming a right-handed double helix.  The two DNA strand are antiparallel, that is, their 5’3’ orientation runs in opposite directions.  The base of both chains lie perpendicular to the axis, and they are stacked on one another.  The nitrogenous bases of opposite chains are paired as the result of BSML-3225 PRELIM CHARGAFF’S RULES  Each complete turn of helix is 34A° long.  The double helix has a diameter of 20A°.  adenine (A) residues = thymine (T) residues guanine (G) residues = cytosine (C) residues.  The sum of the purines equal to the sum of pyrimidine.  The percentage of (G+C) is not necessarily equal the percentage BSML-3225 PRELIM ALTERNATIVE FORMS OF DNA  “B-DNA”  “A-DNA” BSML-3225 PRELIM DNA REPLICATION BSML-3225 PRELIM DNA REPLICATION BSML-3225 PRELIM THE PROCESS elongati terminat initiation on ion BSML-3225 PRELIM BSML-3225 PRELIM BSML-3225 PRELIM BSML-3225 PRELIM BSML-3225 PRELIM BSML-3225 PRELIM PROTEINS REQUIRED IN DNA REPLICATION BSML-3225 PRELIM  DNA gyrase BSML-3225 PRELIM Primase BSML-3225 PRELIM  DNA POLYMERAS E BSML-3225 PRELIM BSML-3225 PRELIM  DNA Ligase BSML-3225 PRELIM THE REPLISOME DNA polymerase III holoenzyme Responsible for the coupling of DNA replication Composed of:  DNA polymerase  sliding clamp  clamp loader BSML-3225 PRELIM BSML-3225 PRELIM REPLICATION PATTERN & TERMINATION BSML-3225 PRELIM opoisomerases BSML-3225 PRELIM REPLICATION AT THE TELOMERE G-rich strand - always at the 3’ end TTGGGG (protozoans) TTAGGG (vertebrates) Telomerase BSML-3225 PRELIM BSML-3225 PRELIM ENZYMES THAT METABOLIZE DNA BSML-3225 PRELIM RESTRICTION ENZYMES endonucleases that recognize specific base sequences and break or restrict the DNA polymer at the sugar- phosphate backbone BSML-3225 PRELIM BSML-3225 PRELIM RESTRICTION ENZYMES Uses: analysis of gene rearrangements Mutation detection DNA recombination in vitro mapping a DNA fragment BSML-3225 PRELIM DNA LIGASE catalyzes the formation of a phosphodiester bond between adjacent 3 ′ -hydroxyl and 5 ′ -phosphoryl nucleotide ends BSML-3225 PRELIM NUCLEASES (EXONUCLEASE) Degrade DNA from free 3 ′ -hydroxyl or 5 ′ -phosphate ends. Use: DNA manipulation in vitro BSML-3225 PRELIM HELICASE AND TOPOISOMERASE Helicase: unwinds dsDNA (breaks hydrogen bonds) Topoisomerase: relieves the tension created by helicase (breaks the phosphodiester linkages in the DNA backbone) BSML-3225 PRELIM METHYLTRANSFERASES catalyze the addition of methyl groups to nitrogen bases, usually adenines and cytosines in DNA strands BSML-3225 PRELIM PHYSICAL PROPERTIES of DNA BSML-3225 PRELIM DNA DENATURATION a process of separating dsDNA into single strands Opposite: renaturation BSML-3225 PRELIM DNA DENATURATION Facto Temperature (high) rs (in vitro) Salt concentration (low) pH (high) BSML-3225 PRELIM MELTING TEMPERATURE (TM) The midpoint of the temperature range over which the DNA strands are half denatured BSML-3225 PRELIM SECONDARY STRUCTURE OF DNA Unusual secondary structures, such as slipped structures, cruciform, and triple- helix DNA BSML-3225 PRELIM TERTIARY STRUCTURE OF DNA Supercoiling BSML-3225 PRELIM BSML-3225 PRELIM End of Part 1  RNA Ribonucleic acid BSML-3225 PRELIM BSML-3225 PRELIM CLASSES OF RNA rRNA tRNA mRNA structural carries amino carry genetic components of acids to the information from ribosome during the DNA of the ribosome translation gene BSML-3225 PRELIM SECONDARY STRUCTURE OF RNA BSML-3225 PRELIM TERTIARY STRUCTURE OF RNA A typical example of RNA pseudoknot motif. S1 and S2 represent double helical stem regions. L1 and L2 are single-stranded loops. BSML-3225 PRELIM RNA TRANSCRIPTION BSML-3225 PRELIM GENERAL FEATURES OF RNA SYNTHESIS BSML-3225 PRELIM PROMOTERS DNA sequences that provide signal for RNA polymerase where RNA polymerase binds 5’ side of the coding strand and to the 3’ side of the template strand BSML-3225 PRELIM RNA POLYMERASE Holoenzyme Sigma subunit core enzyme BSML-3225 PRELIM PROCESS elongati terminat initiation on ion BSML-3225 PRELIM INITIATION BSML-3225 PRELIM ELONGATION BSML-3225 PRELIM TERMINATION BSML-3225 PRELIM TERMINATION BSML-3225 PRELIM OPERONS genes that encode enzymes of certain metabolic pathways close together and under the control of a common promoter Inducer - induction polycistronic message BSML-3225 PRELIM ENHANCERS elements that stimulate transcription cis-acting DNA elements that are not strictly part of the promoter BSML-3225 PRELIM TRANSCRIPTION ATTENUATION a regulatory mechanism that causes premature termination of transcription under certain conditions, thereby preventing the expression of the mRNA required for expression of the corresponding gene products BSML-3225 PRELIM RIBOSWITCHES a region, usually in the 5’-untranslated region (5’-UTR) of an mRNA contains two modules Aptamer Expression platform BSML-3225 PRELIM TRANSCRIPTION IN EUKARYOTES BSML-3225 PRELIM EUKARYOTIC RNA POLYMERASE RNApolymerase I RNApolymerase II RNApolymerase III BSML-3225 PRELIM PROMOTER STRUCTURE Class II promoters Core promoter Proximal promoter BSML-3225 PRELIM PROMOTER STRUCTURE Class I promoters Core promoter element upstream control element (UCE) BSML-3225 PRELIM PROMOTER STRUCTURE Class III promoters nonclassical and it resembles those of Class II genes BSML-3225 PRELIM TRANSCRIPTION FACTORS any protein that regulates transcription but itself is not a subunit of RNA polymerase general transcription factors gene-specific transcription factors (activators) DNA-binding domain activation domain BSML-3225 PRELIM BSML-3225 PRELIM CAP AT THE 5’ END a guanylate residue that is methylated at the N-7 position protect the mRNA from exonucleases degradation BSML-3225 PRELIM POLY-A TAIL AT 3’ END added before the mRNA leaves the nucleus protects the mRNA from nucleases and phosphatases BSML-3225 PRELIM SPLICING cleavage at the 5’ end 3’ end of introns and the joining of the two ends Alternative splicing BSML-3225 PRELIM OTHER RNA- METABOLIZING ENZYMES BSML-3225 PRELIM RIBONUCLEASES degrade RNA in a manner similar to the degradation of DNA by deoxyribonucleases BSML-3225 PRELIM BSML-3225 PRELIM RNA HELICASES catalyze the unwinding of dsRNA BSML-3225 PRELIM NONCODING RNAs BSML-3225 PRELIM NONCODING RNAS (NCRNA) long nc RNAs, such as lncRNA small ncRNAs, such as microRNAs (miRNAs), small interfering RNAs (siRNAs), small nucleolar RNAs (snoRNAs), small nuclear RNAs (snRNAs) BSML-3225 PRELIM PLASMID BSML-3225 PRELIM BSML-3225 PRELIM PLASMID Large plasmid F factor R plasmid Small plasmid BSML-3225 PRELIM BSML-3225 PRELIM