ExAM

EF-Tu

GTPase that transports charged aminoacyl tRNA to A site

EF-G

Hydrolyzes GTP after peptide bond formation, conformation change in EF-G causes movement of small subunit relative to large subunit

Aminoacyl tRNA synthetase

charges tRNAs, is specific to each amino acid

Translation is dependent on what energy source

GTP

Shine-Dalgarno sequence

base pairs with some of small subunit, allows for internal ribosome initiation, allows for translation of polycistronic mRNAs, made of purines, allow for being polycistronic

Initiation

starts with AUG and Shine-Dalgarno sequence ( if bacteria ), Ifs bind to small subunit and attract mRNA, charged fmet tRNA binds to AUG codon of mRNA in P site, forming initiation complex

Elongation

second charged tRNA enters A site facilitated by EF-Tu, peptide bond forms between AAs facilitated by EF-G, uncharged tRNA then moves to E site and out of ribosome

Termination

signaled by UAG, UAA, UGA. GTP dependent release factors cleave polypetide chain from tRNA and releases it from the translation complex

Polysomes

mRNAs with several ribosomes translating at once

Closed loop translation

mRNAs forms loop that is closed by Poly-A binding proteins bind to the cap binding proteins, allows for regulators of translation after stop codon

Beadle and Tatum Neurospora experiment

nutritional requirements were specific and single gened, layed groundwork for Srb and Horowitz

secondary

folding into alpha helix and beta pleated sheet, caused by hydrogen bonding along alpha carbon backbone

tertiary

based on R groups; stabilizing factors: covalent disulfide bonds between close cytosine residues, ionic bonds, hydrophilic R on surface and hydrophobic R on interior, Van der Waals interactions

Post transcriptional modification

N-terminus amino acid removed or modified, individual AA modified (phosphate addition by kinases), polypeptide cleaved to make active form

Somatic mutation

any cell but germ

Germ-line mutation

in gametes and are inherited

Autosomal mutation

within genes on autosomes

X and Y linked mutations

occur on X or Y chromosomes

Point mutations

base substitutions in which one base is altered

Frameshift mutations

result from insertions or deletions of a base pair

Missense mutations

change codon resulting in an altered AA within protein coding portion

Nonsense

changes codon into stop codon

Silent mutation

alters codon but does not result in change in aa, or occurs in non coding portion

Purine

2 ringed base of nucleotide, A and G

Pyrimidine

single ringed base of nucleotide, C, T, and U

Bonding patterns of bases

A with T, C with G, U replaces T

Phenotypic effects of mutation

loss of function, gain of function, morphological, nutritional (biochemical), behavioral, regulatory, lethal, conditional, neutral

Mechanism of DNA errors

tautomeric shifts, slippage, depurination, deamination, oxidative damage, transposons, base analogs, alkylating agents

Tautomeric shifts

transition, alternate forms by single proton shift, Normal: keto T binds to amino A, amino C binds to keto G, Shift: enol T binds to keto G, imino C binds to amino C

Slippage

DNA pol slips or stutters during replication, typically in repetitive sequences, can lead to insertions and deletions

depurination

loss of base

deamination

conversion of amino to a keto group, common to cytosine to change to uracil

keto

T and G, shift to enol

amino

A and C, shift to imine

oxidative damage

superoxide O2., hydroxyl radical OH., hydrogen peroxide H2O2

transposons

integrated into new genomic locations can act as mutagens; jumping genes

Base analogs

substitute for purines/pyrimidines during replication, effect tautomeric equilibriums, 5-BU can be substituted

alkylating agents

donate alkyl group to amino or keto group in nucleotides to later base pairing affinity, EMS

acridine dyes

cause frameshift mutations by intercalating in between the bases

UV radiation

creates pyrimidine dimers (T usually) that distort DNA conformation in way that causes errors in DNA replication

ionizing radiation

X ray, gamma, cosmic - mutagenic

DNA error repair systems

proofreading, mismatch repair, post replication repair, SOS system, photoreactivation repair, Bas and nucleotide excision repair, DNA double strand break repair

Proofreading

exonuclease activity increases fidelity

Mismatch repair

fixes some mistakes that escapes proofreading

How does repair system know which is correct strand?

Methylation

Post replication repair

requires recombination mediated by RecA protein

SOS system

activated by presence of large number of mismatches and gaps, mutagenic but saves cells from death

Photoreactivation repair

nonhuman, removes thmine dimers caused by UV light, dependent on photoreactivation enzyme

Base and Nucleotide excision repair steps

remove mutation with nuclease, gaps filled with DNA Pol, ligation

Base excision repair

single base fix, AP endonuclease , cytosine deamination fixing

Nucleotide excision repair

repairs bulky lesions and involves uvr genes, human alternative to photoreactivation repair

DNA double strand break repair types

homologous recombination repair and non homologous end joining

Homologous recombination repair

fixes double strand break by digesting back the 5’ end of broken helix, this lets undamaged 3’ end of sister chromatid to overhang and allow DNA pol to copy sequence onto damaged strand

Non homologous end joining

activated in G1, error prone, 3 proteins bind to ends, trim ends, and ligate them

Transposable elements

mobile genetic element

insertion sequences (IS)

move from one location to another, can cause mutations

bacterial transposons

larger than IS, can introduce multiple drug resistance to bacterial plasmids, move from plasmids to bacteria chromosomes

autonomous

can move independently

nonautonomous

can only move in presence of autonomous element

Retrotransposons

copy paste, RNA intermediate, can be aut or nonaut

Human transposable elements

SINES and LINES

SINES

nonautonomous, short, cut and paste

LINES

autonomous, long, cut and paste

Cut and paste

DNA

Constitutive enzymes

continuously produced regardless of environment

Operator

region where repressor binds to regulate transcription

Promoter

region where RNA polymerase binds and begins transcription

Repressor

binds to operator to regulate transcription in absence of lactose

Leader

Allosteric repressor

undergoes conformational changes

I-

repressor can not bind to operator

Oc

operator will not bind with normal repressor

Is mutant

operon is superrepressed because inducer can not bind to repressor

I+

does not relieve repression

CAP

positive control of lac operon, absence of glucose cause cAMP to rise and bind to CAP which binds to promoter allowing RNA pol to bind and transcribe

Adenylyl cyclase

converts ATP to cAMP

Trp operon

if tryptophan present, repressed, repressor can only bind if tyrptophan is present

Attenuation

transcription of leader sequence occurs if repressed, termination hairpin forms in the presence of tryptophan ribosome doesn’t stall

Riboswitches

alternative form of mRNA binds with small ligands creating terminator structure

Aptamer

binds to ligand

Expression platform

capable of forming terminator structure

Bacterial small noncoding RNAs

can be negative and positive regulators

Histone modification

functional group adding to N-terminal tails which changes histone affinity, acetylation, methylation, phosphorylation

Chromatin remodeling

repositioning or removal of nucleosomes on DNA

SWI/SNF

loosens attachment of histones, nucleosome core

DNA methylation

decreased gene expression, occurs on CG doublets

Enhancers

enhance transcription, cis acting

Insulators

between enhancer and promoter, allow interactions and block others, cis acting

Silencers

repress the level of transcription initiation, cis acting

GAL

positive control, galactose present, transcription occurs

Junk DNA

does not equal non coding DNA, fraction of human genome that can be mutated without effect

Splicing enhancers and silencers

cis acting, regulate alt splicing

SR proteins

bind to splicing enhancers and activate splicing