Bio112 Unit 2 part 1

4 strucuture levels of chromatin

1- nucleosome (string on a bead) 11 nm

2- chromatin fiber (packed nucleosomes) 30 nm (ZIG ZAG)

3- looped domains (further folding) 700 nm

4- miotic chromosome. 1400 nm

Chromatin

bead-string structure alongside all other interacting proteins (linked by linker DNA)

Histones are high in which animo acids and which histone histone components are conserved

Lysine and Arginine the charged ones, H3, H4

Nucleosome Assembly

(2)H3/(2)H4 bind together to form a tetramer

two H2A/H2B dimers form

Both combine to form a histone octomer

Dyad axis - two-fold axis of symmetry

Histone N-terminal tails are…

long, but no secondary structures (a-helix, b-sheets)

are not required for the formation of core particle

collectively play essential roles in higher oder cjhromatin structure/function

Histone modifiction are usally toward

the N terminal tails (sometimes globular domains)

the lysines in histones can be methylated and acetylated

HAT (Histone acetyl transferase)

Transfers acetyl group onto histones allowing for greater gene expression by making it less possible (acetyl group removes lysines positive charge that wraps the negative DNA tightly)

(Recruited by activators and repressors as coactivators and corepressors)

Discovered by U of R scientist David Allis

HDAC (Deacetyl-transferase)

Removes acetyl groups (discovered by stuart schreiber)

(Recruited by activators and repressors as coactivators and corepressors)

Methylation affects…

transcription (up or down depending on H3-K4 up or H3-K9 down) and repair

Acetylation affects…

Transcription, Repair, replication, and condensation.

Writers and erasers

introduces the histone modifications (HAT)

Remvoes histone modifications (HDAC)

Readers

Binds to a modified histone- Recognizes modificaitons and translates it into outcomes

could activate 

1. ATP depdendent Chromatin remodeling Complex (SWI/SNF→gene regulation)

2. Histone modifier (reader/writer)

3. Factor that syabilizes higher order chromatin structure 

4. Another gene regulator

Histone code Hypothesis

The modification state of a histone is associated with a specific protein that results in a certain function.

(EX. Acetylatation →Bromodomain leads to transcription)

Eukaryotic gene regulation (states)

silent state - hypoacetylation (h3,h4) hypomethylation of H3

ground state - H3, H4 acetylation, H3 methylation 

poised state/active state - Tri methylation H3, Hyperacetylation of H3, H4

Modifications of chromatin 

Sliding, ejecting, unrapping, deposition, assembly. 

Chromatin remodelers form remodeling complexes like 

SWI/SNF

PHO8 example of chromatin in gene regulation

Phosphate depletion→PHO4 activator to bind to gene→HAT(SAGA) binds to PHO4 and temporarily hyperacetyl neurby nucleosomes→

SWI/SNF binds to gene and ejects histone→RNA pol II binds→gene activation

Cell cycle (HO yeast) example of chromatin in gene regulation

Anaphase has SWI5 → Telophase chromatin remodeler then HAT → SBF 

(Recurit of translation activators is gene specfic is the main point)

Euchromatin vs Heterochromatin

Euchromatin is less condensed, hyperacetylated histones, accessible to factors trhat allow for gene expression

Heterochromatin - Highly condensed, near centromeres and telomeres, Hypoacetylate dhistones, not accesible to binding factors > inhibits gene expression

Formation of Heterochromatin

1. Recruitment of writer

2. Modification of nearby nucleosome (wraps tighter)

3. Recruitment of reader-wroter complex 

4. Modification of second nucleosome

5. repeat 2-3 and modification of third nucleosome

6. Repeat loop for subsequent chromsomes

Aminoacyl-tRNA function

Matches 20 amino accids to codons in mRNA

(amino acid-tRNA-codon)

Aminoacyl-tRNA synthetase binds the amino acid to the tRNA and recognizes correct tRNA sites

Aminoacyl-tRNA mismatch problem

Ribosomes don’t care about the actual amino acid bounds to the tRNA so there could a AAtRNA with the codon for one amino acid that’s actually carrying another. Example: Val-tRNA^ala

Double sieve mechanism 

how amino tRNA selects the correct amino acid- first sieve filters by size removing all too large amino acids and second lets the too small ones to pass though leaving just the correct amino acid. 

RIbosomes

large and small subunit 

prokaryotes (30s small 50s large)

eukaryotes (40s small 60s large)

Peptidyl transferase

transfers peptides to A site aminoacyl amino acid

Prokaryotic initiation translation

1. Intiator tRNA(fmet) binds to the P site Initiator

2. RBS(Ribosome binding sequence) part of the mRNA transcript is brought over and slides until it reaches it reaches a point at which AUG is lined up with the fMET in the p-site

3. Large subunit binds and the aminoacyl-tRNA bonds to A site with first peptide bond forming.

4. Large subunit trasnlocates one codon, followed by small subunit moving the growing chain to the p site and ejecting the initiator RNA in the E site. 

Eukaryotic initation translation

1. Initiator tRNA(fmet) binds to the P site 

2. Initiator complex recognizes 5’cap mRNA, sliding it until AUG matches with fmet

3. Large subunit binds and aminoacyl-tRNA bonds to A site with first peptide bond forming.

4. Large subunit translocates one codon, followed by small subunit moving the growing chain to the P site and ejecting the initiator RNA in the E site. 

Elongation

1. Aminoacyl tRNA binds to A-site new pepetide bond is formed (Peptidyl transferase transfer peptide to new amino acid, new amino(amine group) acid acts as nucleophile donating e-

2. translocation of large subunit and then small subunit leading to ejection of previous tRNA

3. Elongation factors drive translation foward

Termination

1. Ribosome reaches a stop codon (UAA, UAG, and UGA)

2. Release factor binds to A site (looks like tRNA

3. peptide gets transfered to a water leading ot hydrolysis and the end of protein synthesis

4. mRNA is reease →two subunits dissociate

Translation inhibitors

Acts as an Antibiotic

Examples (Tetracycline, streptomycin, Chloramphenicol, etc)