Prokaryotic Gene Expression Flashcards
what are the proteins in a prokaryotic RNA pol?
beta, beta’, alpha 1 and 2, and omega–> sigma is added for specificity, but the pol will work without it
RNA pol in proks start to transcribe from NICKED DNA
ya
RNApol has 5’ to 3’ pol and helicase activity
ya
what is the purpose of the omega unit?
helps assembly
what are the alpha units required for?
required for assembly, have some DNA specificity
what are the beta subunits required for?
catalytic proteins for RNA polymerization
assembly order for RNA pol
alpha dimerizes, binds beta, omega and beta’ bind, then both complexes bind together–sigma comes in later after binding to DNA
different sigma factors for different genes promoters–> sigma 70 is common for many genes though
ya
sigma has 4 conserved domains–> domains 2 and 4 contact the DNA t the positions -10 and -35, respectively
ya
the closer the consensus seq is for sigma tobind to, the more increased the probablility is that RNA pol will bind there
ya
-10 box is the Pribnow box–TATA boxes are eukaryotic
aya
additional binding sites for RNA pol
-40-60, UP element and extended -10, TG seq at -14/-15
what are the purpose of Gre A and B
enhance hydrolytic removal–>RNA pol backtracks 3-10 nt and cuts by endonuclease activity–involved in proofreading of RNA pol
what are the proofreading activities of RNA?
pryphosphorolytic rxn–cuts two phosphates off a triphosphate nt; hydrolytic removal–>RNA pol backtracks 3-10 bases and cuts via endonuc activity
termination in bacteria
2 types: rho depedent and rho independent
what is rho independent termination?
utlized GC-rich inverted repeat seq; has no effect on DNA but causes a stem-loop form from teh RNA; stem forms and yanks RNA rom the pol; need a stretch of ~3 A:U pairs so binding will be weak enough to free the RNA
what is rho dependent termination?
used less often that rho independent; rho is a hexameric protein that binds the RNA t the rut site (rho utilization); rho binds RNA & yanks it out; • Rho-dependent might help to make it more error proof
Rut site can only be bound after translational stop site–otherwise ribosomes would constantly be feeding on this and disrupting Rho from reaching the site
the rut site is high in Cs
ya
translation starts before transcription is finished
ya
what are polycistronic genes?
several genes are transcribed to one mRNA, but one mRNA makes many proteins–> generally, the expression of several genes at once is useful because they will normally be needed to be used together
describe the 70S ribosome
2 subunits: 50S and 30S; 50S contains the 23 and 5 S and 34 proteins; 30S contains the 16 S rRNA and 21 protiens
what role does the 16S rRNA play?
ensures accurate tRNA binding and placement of translational initiation (through hoogsteen binding)
what role does the 23S rRNA play?
produces peptide bonds between amino acids and the growing protein
what role does the 5S rRNA play?
structural integrity
what are the initiation factors and what do they do?
IF-3 binds to the 30S subunit and prevent association with the 50s for the time being; IF 1+2 come in and bind the 30S (IF 2 has fMet on it) at the S-D seq; IF 1+3 are lost so now the 50S subunit can bind–> fMet now in P site; IF2 is released from the A site–>free A-site can now bind a tRNA
describe synthesis of a protein
the 23S rRNA catalyzes the transfer of the growing polypeptide to the newest aa; ribosome slides forward one codon–>newest tRNA enters P site from A site, and the tRNA in the P site goes to teh E site–>now a new tRNA can enter the P site
describe elongation in proks
aminoacyl-tRNA in brought to A site by EF-Tu in a GTP-hydrolyzing step; 23S ribosomal RNA transfers the growing peptide to teh N-terminus of the new aa; EF-G ratchets the transition-state ribo forward so that A-site tRNA with the protein in placed into the P ist
speed of elongation?
60 nt/sec or 20 aa/sec
describe translation termination in proks
no tRNA binds to a stop codon; instead release factors 1 or 2 (depending on the stop codon) binds to the stop codon and 23S rRNA hydrolyzes the bond bw tRNA and protein; RF-3 then dissembles the ribosome
