Wednesday, April 16, 2014

Biochem Mock Board Review #3!

Molecular Biology and Epigenetics
I.

DNA and Epigenetics!

A. Chromatin!
1. DNA + Histones!
a) Histones !
(1) have 8 parts - two of each of: H2A, H2B, H3, H4!
(2) H1 clips DNA around Histone!
(3) Histones have + charge from Lys and Arg amino acids!
b) DNA - negatively charged so its attracted to the + charge on histones!
2. Euchromatin!
a) Loose chromatin that can be transcribed (read) to RNA!
b) ACETYLATION = less positive charge = euchromatin!
3. Heterochromatin!
a) Condensed/tight chromatin that is not able to be transcribed to RNA!
b) METHYLATION of histones and/or DNA can lead to condensed chromatin!
(1) DNA methylation occurs on cytosine (C) that is next to guanine (G) aka
CpG islands!
B. DNA & RNA!
1. Nucleotides!
a) Bases!
(1) Purines (A & G) - Pure As Gold - 2 rings !
(a) When you make purines you start with the sugar and add the base!
(2) Pyrimidines (C, T, U) - CUT the Py (pie) - 1 ring!
(a) When you make pyrimidines you start with the base (orotic acid)
and add the sugar!
(b) U is only in RNA!
(c) T is only in DNA - must use Thymidylate Synthase to get dTMP!!!

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b) Sugars!
(1) RNA - ribose!
(2) DNA - deoxyribose!
(3) You make the ribose (RNA) versions first!
(4) ribose can be converted to deoxyribose by ribonucleotide reductase!
c) Phosphate - linked by 3 - 5 phosphodiester bond!

d) Notice Carbamoyl Phosphate in the figure above!

(1) This intermediate is in urea cycle too!
(2) If the urea cycle is defective at ornithine transcarbamoylase, you have
excess carbamoyl phosphate that gets converted to OROTIC ACID!
(3) This causes OROTIC ACIDURIA, but if you give them uridine, they are
ok!
C. Genetic Code!
1. Definitions!
a) Unambiguous - a codon codes for only 1 AA!
b) Degenerate/Redundant - more than 1 codon gives the same AA!
c) Nonoverlapping - you read DNA from point A to point B without skipping!
d) Universal - all organisms have the same genetic codons!
e) Start codon = AUG, Stop Codons = UGA, UAA, UAG!

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2. Mutation Types - in order from best to worst!
a) Silent - the codon changes, but you get the same AA!
b) Missense - the codon changes, the AA changes, but protein structure/
function is preserved!
c) Nonsense - STOP CODON! - Stop the Nonsense!
d) Frame Shift - you skip a part of the DNA so everything is off by 1 (or 2,
etc). Usually causes nonfunctional protein!
II. DNA Replication & Repair!

A. Replication - OCCURS 5 TO 3!
1. Origin of replication - wherever DNA replication begins!
a) Prokaryotes only have 1!
b) Eukaryotes can have a bunch at once!
2. Replication fork - the place where the replication is happening!
3. Helicase - unwinds!
4. Single Stranded Binding Proteins - keeps unwound DNA from getting tangled!
5. Topoisomerases - untwist DNA (relieve supercoils)!
6. Primase - makes primers so DNA polymerase has somewhere to start!
7. DNA Polymerases - they do the work!

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a) DNA pol 3 - adds 5 > 3 adding power and 3 > 5 correcting
(exonuclease) power!
b) DNA pol 1 - removes RNA primers with 5 > 3 exonuclease !
8. DNA Ligase - fills in any gaps!
9. Telomerase - adds DNA to 3 end to keep from losing the DNA ends every
time you make a copy!
B. DNA Repair!
1. Nucleotide Excision Repair - removes and replaces base, sugar and
phosphate when nucleotide is damaged!
2. Base Excision Repair - only removes a damaged base and replaces!
a) Uses AP Endonuclease!
3. Mismatch repair - when you replicate DNA, this makes sure the bases match
up right!
III. Transcription - making mRNA!
A. Regulators!
1. Promoter - where RNA polymerase and Transcription Factors bind!
2. Enhancer - where activating Transcription Factors can bind!
3. Silencer - where inhibiting Transcription Factors can bind!
B. RNA Polymerase - make RNA!
1. Remember r.m.t.!
a) rRNA by RNA pol #1!
b) mRNA by RNA pol #2!
c) tRNA by RNA pol #3!
C. Processing mRNA!
1. Cap on 5!
2. Poly-A tail on 3!
3. Splice out introns!
a) spliceosome and snRNPs!
b) Just remember the word lariat!
IV. Translation - making protein (N terminus to C terminus)!

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A. Uses ribosome !
1. Uses the A, P and E sites in that order (ribosomes go APE when they get to
make proteins!)!
a) Initiation - get the ribosome, rRNA, mRNA and tRNA all together!
b) Elongation - recruit tRNAs and AA and form the bonds!
c) Termination - STOP codon!
2. tRNAs bring AA to the ribosome!
a) Uses anticodon to read mRNA!
b) 3 end of tRNA binds the AA!
c) Aminoacyl-tRNA synthetase makes sure the tRNA has the right AA!
V. Important Molecular Biology Techniques!
A. PCR - polymerase chain reaction - making copies of DNA pieces!
1. Denature DNA by heating!
2. Anneal by cooling DNA down in presence of primers!
3. Elongate by letting DNA polymerase make copies using primers!
B. The Blots!
1. Remember SNoW DRoP!
2. Southern Blot detects DNA!
3. Northern Blot detects RNA!
4. Western Blot detects Protein!
C. ELISA - Enzyme linked immunosorbent assay!
1. Take something you want to detect (antigen) in a sample !
2. Add antibodies against that antigen to the sample!
3. Add a peroxidase enzyme to make color change if antigen is present