(lanes in right hand figure of gel: RNA polymerase is present (+) or absent (-).
RNA polymerase binding is suggested in the +2 to -35 and -39 to -54 regions.
C is a control where the DNA was cleaved chemically)
Transcription Initiation and Control.
DNA footprinting methodology:
(lanes in right hand figure of gel: RNA polymerase is present (+) or absent
(-).
RNA polymerase binding is suggested in the +2 to -35 and -39 to -54
regions.
C is a control where the DNA was cleaved chemically)
Another DNA footprint for RNA polymerase.
Analysis spans a larger range of base pairs than above. Sequencing lanes
by Maxam-Gilbert Sequencing
Lac operon transcription control region, as
determined by DNAse I footprinting analysis.
Prokaryotic RNA polymerase binding to the
promoter
Comparison of prokaryotic strong promoters
for s70
Lac repressor footprint at the operator
DNA binding proteins:
Homodimers (or
homotetramers (lac repressor is a dimer of dimers)) that contain structural
motifs for DNA binding.
General categories of structural motifs for
proteins that bind to DNA in a regulatory capacity:
1. Helix-turn-helix (or helix-loop-helix) (turn is a b-turn,
loop is a random loop)
Helix-loop-helix in the lac repressor:
2. Zinc-fingers (red ball is zinc ion)
3. Leucine Zipper - attached to a DNA
binding region
Other DNA binding protein motifs: homeodomain (binds Homeobox DNA sequence)
Positive regulation of the lac operon
(activation) is through cAMP-CAP (CRP)
homodimer, red in center is cAMP, yellow is region that
interacts with RNA pol
Catabolite repression of lac operon
prevents expression when glucose is present regardless of the presence of
lactose.
CRP and RNA pol are cooperative in their
binding to the promoter.
Multi-level regulation in the Tryptophan
operon (Trp operon):
1. Repression by Trp repressor (Trp is the effector)
2. Transcription attenuation (links transcription and
translation)
(we'll
discuss this more in chapter 11)
Trp repressor dimer bound to Trp (red)
One protein can act as a repressor or
activator depending on the presence of an effector molecule in the ara operon:
some genes contain promoters that are
recognized by other sigma factors:
s54
activators bind far upstream from the start site, and can act even when their
binding sites (enhancers) are more than 1kb away:
NtrC is a s54 activator
for the glutamine synthetase (glnA) gene:
Physical evidence for above model.
