BCHM 3050 1ST Edition Lecture 15 Outline of Last Lecture I. Levels of Structure in DNAII. Formation of Phosphodiester BondsIII. B-DNA Spatial DimensionsIV. Stabilizing Forces in DNAV. Functions of Major GroovesVI. Functions of Minor GroovesVII. DNA Structural VariationsOutline of Current LectureI. Super-Coiling of DNAII. Packaging of Eukaryotic DNAIII. RNA vs. DNAIV. Types of RNAV. Transfer RNAVI. Ribosomal RNAVII. Griffith’s ExperimentVIII. Avery-MacLeod-McCarty ExperimentIX. Griffith and Avery’s ExperimentX. Hershey and Chase ExperimentCurrent LectureThese notes represent a detailed interpretation of the professor’s lecture. GradeBuddy is best used as a supplement to your own notes, not as a substitute.I. Super-Coiling of DNAa. A and T have a double bond between themb. G and C have a triple bond between themc. No two people have the same sequence of bases (except identical twins)d. DNA undergoes changes over time – epigeneticse. DNA has to be supercoiled in order to fit in such a compact spacef. Supercoiling makes it inaccessible which is a way the cell protects the DNA from mutagensg. Supercoiling makes sure that the replication factors (needed for the DNA replication process) access the DNA only at specific timesh. Higher organisms have maximum supercoilingII. Packaging of Eukaryotic DNAa. The dots on the print out are groups of proteins called histones that sit on the DNAb. Nucleosome – group of 8 histone proteins that bind to the DNA at certain intervalsc. When cell is splitting, DNA is at its maximum supercoiling à tightly packed and not accessible to anything elsed. Heterochromatini. Nucleosome are so tightly packed that there is no DNA between ii. highly supercoiled state; cannot participate in anythinge. Euchromatini. Certain enzyme push the histones apart and expose DNA between ii. less super coiling; “beads in a string”; state when replication and other processes occuriii. DNA is exposed and can get attacked by other moleculesf. 140 base pairs of DNA rapped around nucleosomeg. Linker DNA – 60 base pairsh. 140 more base pairsi. H1 does not have a role in the part of the nucleosomej. Overall charge of DNA is negative – which means the histone proteins have to be positively charged; basic amino acids (especially arginines and lysines that get chemically)k. Modifying the arginines and lysines in histones changed it from heterochromatin to euchromatinIII. RNA vs. DNAa. RNA is much more abundant than DNA because there are many more copies of RNA’s (messages, transfer species, ribosome numbers) made than the 2 copies ofDNA.b. Modified bases in tRNA include pseudouridine, dihydrouridine, 4-thiouridine, 1&2-methylguanosines especially in loops.c. U is replaced by T (i.e. methylated uracil), because T will only base-pair with A; U,however is the only base that can base pair with any of the other bases. Evolutionarily, this could not be “tolerated” to maintain the fidelity required in DNA base pairing.d. RNA is more transient and more easily degradede. Secondary structure of RNAi. Single strand of RNA is very flimsyii. Can make transient bonds with its own strandIV. Types of RNAa. snRNA’s thought to complex with proteins to form snRNPs (small nuclear ribonucleo proteins), are involved in removing introns and then splicing exons together to form functional mRNA’s.b. snoRNA’s (small nucleolar) –snoRNA - small nucleolar RNA, forms snoRNPs, which process rRNA, mostly by methylation and isomerisation; thought to be involved in ribosome biogenesis.c. siRNA - small interfering RNA, involved in gene silencing and regulation. d. gRNA - guide RNA, needed for RNA editing, the removal and insertion of bases into mRNA. e. tmRNA - an RNA molecule that disengages ribosomes from stalled translation of mRNA in bacteria. f. telomerase RNA - an RNA molecule that forms much of the structure and all of the template required by telomerase. g. hnRNA - rag-bag of unprocessed pre-mRNA transcripts and other heterogeneous nuclear RNAs of less well defined function. h. tRNA and mRNA – involve in translationi. snRNA – involved in splicingV. Transfer RNAa. tRNA stays in the clover leave position because of the modifications of uracil and guanineb. Know the names of the modifications of uracil and guanine found in tRNAc. Amino acid can only attack to the tRNA at the 3’ end (anticodon end)d. tRNA participates in translatione. All tRNAs have CCA followed by an amino acid at the 3’ endf. Each amino acid has its own special tRNAVI. Ribosomal RNAa. rRNA lives in the ribosome b. Very rigid and assume a specific structurec. rRNA sits on the ribosome in this specific conformation in all organismsd. rRNA plays important role in translation and protein synthesisVII. Griffith’s Experimenta. Isolated 2 different strains of the bacteria – R and Sb. R = “rough” strain is devoid of capsule, which is what infects; this is avirulent and allowed the mice to livec. S = “smooth” strain with capsule; virulent and kills miced. Experiment:e. heat kills the S strain and infected the mouse à mouse lived (so heat can destroy whatever is making this bacteria infectious)f. Combined R strain and the heat killed S strain and injected into mouse àmouse died; strain of virulentg. Conclusion – somehow the R strain was transformed in the presence of heat killed S strain to make it virulenth. Some unknown particle is a transforming agent (which we know today is DNA)i. R bacteria + heat killed S = virulent R strainVIII. Avery-MacLeod-McCarty Experimenta. Knew we could isolated DNA, protein, carbohydrates, lipidsb. Avery-MacLeod-McCarty – hypothesized that one of these 4 molecules was the transforming agentc. Amylase kills all carbohydratesà heat killed S and R combo still killed mice and transferred Rd. Protease kills proteins à still transformed Re. Dnase degraded DNA à heat killed S could not transform R so this shows that the DNA is the transforming agentf. RNase degraded RNA à transformed Rg. DNA could also transform over generations and between one species and anotherIX. Griffith and Avery’s Experimenta. Non-virulent strain injected in mice, live.b. Virulent strain injected in mice, die.c. Heat killed virulent strain injected in mice, live.d. Heat killed virulent strain and non-virulent strain injected in mice, die.X. Hershey and Chase Experimenta. Proved that DNA is a genetic materialb. They used a virus and bacteriac. Virus injects something inside of the host cell
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