Bio 99 Final Review

What is the main purpose of regulation?

Transcription initiation

Where does RNA polymerase bind?

Promoter

Is there a primer for transcription?

Naur

How is a consensus sequence confirmed?

1) Sequence conservation

2) Mutation

3) DNA footprinting

What are the common themes of E.coli promoters?

1) -35 box TTGACA (TG element)

2) -10 box TATAAT (Pribnow)

3) Purine start site (A or G)

Why is the -10 box ideal for unwinding?

AT has only 2 hydrogen bonds to break

What is the "scrunching" model?

DNA is pulled into RNA pol for "melting" and begins RNA synthesis

How does rifampicin work?

Binds to ꞵ subunit and blocks initiation

Does elongation have a sigma complex?

Naur

What is the core complex (claw-like, has a channel)?

ɑ2ꞵꞵ', transcribes ssDNA or nicked DNA

What is the holoenzyme?

ɑ2ꞵꞵ'σ (or ɑ2ꞵꞵ'σω), promoter specific

How does GreB rescue an arrested complex?

Binds and provides a second Mg2+

Speed: DNA replication > RNA transcription > translation. Why?

More error-prone processes are slower

How many Rho factors are needed for Rho-dependent termination?

6

What is the name of professor Tsai's dog?

Howie :)

What is the most common process that determines protein concentration in a cell?

Transcription initiation

What are regulators?

DNA-binding proteins that bind to specific sites near promoters and affect the ability/affinity of RNA pol to initiate

What are repressors?

Regulators that inhibit/block binding of RNA pol

What are activators?

Regulators that bind nearby and have affinity for RNA pol

What does negative control involve?

Repressor

What does positive control involve?

Activator

What is the function of galactoside (Y) permease?

Transport lactose into the cell

What is the function of beta-galactosidase (Z)?

Break down lactose into glucose and galactose

What is allolactose/IPTG?

Inducer

What is an operon?

A segment of DNA that consists of the operator, the promoter, and the genes being controlled.

Is lac operon a palindrome?

Yes, sequence 5'AATTG

How many lacI (repressors) bind?

4

What are cis factors?

DNA sequences in the vicinity of genes

What are trans factors?

Diffusible protein factors that bind to cis DNA sequences

What is the phenotype of lacI mutant (i⁻)?

Lac genes expressed continuously

If a second wild type or normal copy of the lacI gene is introduced into the lacI mutant cell, what would be the phenotype of this partial diploid?

Lac genes expressed efficiently only in the presence of lactose

What is the phenotype of a lacI S (super repressor) mutant?

Lac genes never expressed efficiently

What is the phenotype of a lacI S mutant that has a second normal copy of the lacI gene?

Lac genes never expressed efficiently

What is the phenotype of an operator mutant (Oᶜ) that prevents the binding of the repressor?

Lac genes expressed continuously

What is the phenotype of a promoter mutant (lacP) which has a mutation in the promoter of the lac operon?

Lac genes never expressed efficiently

What are catabolic operons and how are they regulated?

Operons devoted to breaking down more complex molecules, regulated by induction

What are anabolic operons and how are they regulated?

Operons devoted to biosynthesis, regulated by repression

Where does Trp bind on DNA?

Major groove

How much of the human genome encodes for proteins?

1% (non-coding regions are for gene regulation)

How many classes of DNA do humans have?

3 (# of rises = # of classes)

What are the roles of the 3 RNA polymerases and their sensitivities to ɑ-amanitin?

1) RNA pol I: makes rRNA, resistant

2) RNA pol II: makes mRNA, very sensitive

3) RNA pol III: makes tRNA, less sensitive

What is TFIID composed of?

TATA binding protein (TBP) and TBP associated factors (TAFs)

Where does TBP bind?

Minor groove

Which promoters does TFIID bind to?

TATA, Inr, DPE

Which promoter does TFIIB bind to?

BRE

What causes xeroderma pigmentosum and Cockayne Syndrome?

Mutation in TFIIH

What makes pol II unique?

It has a C-terminal domain (CTD)

What amino acid is targeted for phosphorylation?

Ser

What enzyme adds phosphate on CTD?

TFIIH

What factors prevent CDT phosphorylation?

Negative elongation factor (NELF) and DRB-sensitivity-inducing factor

What kinase phosphorylates CDT?

P-TEFb

What is the role of fork loop 1 in transcription elongation?

Prevents premature unwinding

What is the role of the rudder in transcription elongation?

Prevents DNA from rebinding to newly synthesized RNA

What is the role of the lid in transcription elongation?

Wedge and guide

After elongation, how is pre-mRNA processed?

Add 5' cap, splicing, cleave poly-A tail

What recruits the capping enzyme (+)?

CTD (-) of RNA pol II due to charge-charge attraction

To target transcript for capping, where is CTD phosphorylated?

Ser-5

How does pre-mRNA produce different mRNAs?

Alternative splicing

What diseases do splicing errors cause?

Muscular dystrophy, Parkinson's, Alzheimer's, cystic fibrosis, cancer

What conducts splicing?

Spliceosome (made up of snRNA)

What is heterochromatin?

Condensed, transcriptionally inactive, inaccessible DNA

What is euchromatin?

Loose, transcriptionally active, accessible DNA

What are the 3 eukaryotic gene repressor functions?

1) Competitive DNA binding

2) Masking activation surface

3) Direct interaction with GTFs

What does the mediator complex interact with?

CTD of pol II

What are the main amino acids in histone?

Lysine (+) and arginine (+)

What proteins are involved in deacetylation?

Sir2, Sir3, Sir4

What is the function of the chromodomain?

Allows proteins (HP1 in methylation) to bind to methylated histones

What is the function of the chromoshadow domain?

Allows HP1 (involved in methylation) to bind to other HP1 proteins

What is the function of SWI/SNF?

Move nucleosomes away from promoter sequences to facilitate transcription

Do microRNAs get a 5' cap and poly-A tail?

Yes

What are the differences between miRNA and siRNA?

1) miRNA is natural, siRNA is artifical

2) miRNA results in translation inhibition, siRNA can result in inhibition or degradation

For CRISPR, what 3 things are added to a human cell?

1) Cas9 (nuclease)

2) Gene specific crRNA

3) tracr RNA (links crRNA to Cas9)

What did Nirenberg and Leder discover?

Filter binding assay: pair codons with amino acids

What caps the first amino acid in bacterial initiation?

Formyl (HC=O)

What does the Shine-Dalgarno (SD) sequence complement?

16S RNA to establish open reading frame

What is the purpose of polysome formation?

Add 5' cap and poly-A tail to stabilize and protect mRNA

What is at the active site of ribosome?

Adenine

Where does puromycin bind?

A site