Lecture 45-58

Question 1

Transcription

Answer 1

Process of RNA synthesis with DNA template. Catalyzed by RNA polymerase. Begins at promoter and ends at terminator

Question 2

Trans-acting factors/elements

Answer 2

Usually DNA-binding proteins (ex: TFs) but include some non-coding RNAs. Diffusible and function at multiple sites

Question 3

Cis-acting factors/elements

Answer 3

Usually DNA sequences that are part of genes

Question 4

mRNA

Answer 4

Encodes primary AA sequence for proteins (template for translation)

Question 5

tRNA

Answer 5

Carries AA into catalytic site of ribosomes. Base pairs to mRNA to ensure correct AA sequence

Question 6

rRNA

Answer 6

Structural components of ribosomes

Question 7

Primary transcript

Answer 7

Finished RNA molecule beginning at promoter and ending at terminator. Used as mRNA in bacteria, but in eukaryotes undergoes further modification

Question 8

Template strand

Answer 8

DNA strand used in transcription to synthesize RNA. Equals the reverse complement of the coding strand and the primary transcript

Question 9

Coding strand

Answer 9

DNA strand that resembles the RNA primary transcript (except T for U)

Question 10

Prokaryotic promoter

Answer 10

Cis-acting element where RNA pol binds to initiate transcription. Have orientation polarity. Consensus sequences on coding strand:

-35: TTGACA
SPACER
-10: TATAAT
SPACER

Most primary transcripts begin with purine and often with CAT sequence (A = +1)

Question 11

Gene

Answer 11

Unit of heredity with some function in the organism.Most encode information for a protein and includes the DNA encoding the protein and regulatory elements needed for transcription

Question 12

ORF (cistron)

Answer 12

Sequence of bases that encodes primary protein sequence

Question 13

Operon

Answer 13

Coordinately regulated gene clusters. May contain multiple ORFs operating with one promoter and terminator. Yields one polycistronic mRNA

E. coli atp operon = 9 ORFs for ATP synthase

Question 14

Constitutive promoter

Answer 14

Promoter that is always active with rate of transcription initiation determined by sequence identity to consensus

Question 15

Strong promoter

Answer 15

Has high sequence identity to promoter consensus. Mutations that move toward consensus increase rate of transcription initiation

Question 16

Weak promoter

Answer 16

Has several base changes from promoter consensus. Mutations that move away from consensus decrease rate of transcription initiation

Question 17

RNA pol (E. coli)

Answer 17

Catalyzes transcription in 5’-3’ direction along template strand with similar mechanism to DNA pol. Primer independent. Holoenzyme responsible for initiation and synthesis of first 10 nt. Holoenzyme = a2BB’wo subunits

Core enzyme carries out elongation.
Core enzyme = a2BB’w subunits

No proof-reading (3’-5’) exonuclease. High error rate (balanced by high RNA turnover rate)

Question 18

o (sigma) subunit

Answer 18

Recognizes promoter. 7 different sigma subunits in E. coli. Sigma 70 binds to most promoters (consensus sequence). Other sigmas bind to different promoter consensus sequences for genes with specialized functions

Question 19

a2 (alpha) subunit

Answer 19

Essential for enzyme assembly and involved with activator interactions

Question 20

B, B’ (beta, beta prime) subunits

Answer 20

Form catalytic core

Question 21

w (omega) subunit

Answer 21

Provides structural stability

Question 22

Transcription initiation

Answer 22

RNA pol is DNA template-dependent but primer-independent. Uses ribonucleotide triphosphates (NTPs) as substrate. Holoenzyme binds promoter (closed complex). Unwinding of 12-15 bp DNA into transcription bubble (open complex). Holoenzyme synthesizes first 10 nt (1 nt/sec)

Question 23

Transcription elongation

Answer 23

Sigma dissociates and is replaced by NusA - allowing RNA pol to complete promoter clearance. Elongation rate of 50-90 nt/sec but can be slowed by secondary RNA structure formation in transcript. Transcription bubble is 12-15 bp and DNA-RNA hybrid is 8 bp. Topoisomerases relieve supercoiling.

Termination results in RNA release and dissociation of core enzyme from DNA.

Question 24

Rho-dependent termination

Answer 24

Requires Rho (p) protein. Rho is hexameric helicase that binds to rut site on nascent RNA. Translocates 5’-3’. When RNA pol pauses due to RNA secondary structure, termination occurs when/if Rho contacts RNA pol

Question 25

Rho-independent termination

Answer 25

Requires termination signal in nascent RNA. Occurs due to formation of stable hairpin followed by series of Us. Hairpin causes only weak AU bp between RNA and DNA. DNA-RNA hybrid is unstable and causes RNA disassociation