Transcription

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Transcription
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MVA Fig. 26.8
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Figure 31-11a X-Ray structure of Taq RNAP core
enzyme. a subunits are yellow and green, b
subunit is cyan, b subunit is pink, w subunit is
gray.
What do you notice about this structure?
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Figure 31-13b Model of the open (Rpo) complex of
Taq RNAP with promoter-containing DNA showing the
transcription bubble and the active site.
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Figure 31-14 The two possible modes of RNA chain
growth. Growth may occur (a) by the addition of
nucleotides to the 3 end and (b) by the addition
of nucleotides to the 5 end.
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How could you distinguish between these two
possibilities?
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Cordycepin triphosphate
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Initiation
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MVA Fig. 26A. DNA footprinting (Nuclease
protection assay)
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DNA Footprinting
http//users.rcn.com/jkimball.ma.ultranet/BiologyP
ages/F/ Footprinting.html
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Figure 31-10 The sense (nontemplate) strand
sequences of selected E. coli promoters.
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Figure 31-12a The sequence of a fork-junction
promoter DNA fragment. Numbers are relative to
the transcription start site, 1.
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MVA Fig. 26.6
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Figure 31-15 RNA chain elongation by RNA
polymerase.
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Figure 31-16 An electron micrograph of three
contiguous ribosomal genes from oocytes of the
salamander Pleurodeles waltl undergoing
transcription.
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MVA Fig. 26.8
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RNA Backtracking
MVA Fig. 26.10
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MVA Fig. 26.15
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MVA Fig. 26.16
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Figure 31-18 A hypothetical strong (efficient) E.
coli terminator.
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Regulation Possiblilites
Regulate transcription Regulate
translation Regulate activity
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The lac operon

• E-coli uses three enzymes to take up and
  metabolize lactose.
• The genes that code for these three enzymes are
  clustered on a single operon the lac Operon.

Whats lactose??
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Figure 31-2 Genetic map of the E. coli lac operon.
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The lac repressor gene

• Prior to these three genes is an operator region
  that is responsible for turning these genes on
  and off.
• When there is not lactose, the gene for the lac
  repressor switches off the operon by binding to
  the operator region.
• A bacteriums prime source of food is glucose.
• So if glucose and lactose are around, the
  bacterium wants to turn off lactose metabolism in
  favor of glucose metabolism.

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Isopropyl thio -? -D- galactoside
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Figure 31-25 The base sequence of the lac
operator.
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• Lac repressor binding to DNA animation
• http//molvis.sdsc.edu/atlas/morphs/lacrep/index.h
  tm

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Figure 31-28a X-Ray structures of CAPcAMP
complexes. (a) CAPcAMP in complex with a
palindromic 30-bp duplex DNA.
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Figure 31-36 X-Ray structure of the lac repressor
subunit.
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Figure 31-37a X-ray structure of the lac
repressor-DNA complex.
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Induction.

• Allolactose is an isomer formed from lactose that
  derepresses the operon by inactivating the
  repressor,
• Thus turning on the enzymes for lactose
  metabolism.

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The lac operon in action.

• When lactose is present, it acts as an inducer of
  the operon (turns it on).
• It enters the cell and binds to the Lac
  repressor, causing a shape change that so the
  repressor falls off.
• Now the RNA polymerase is free to move along the
  DNA and RNA can be made from the three genes.
• Lactose can now be metabolized (broken down).

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When the inducer (lactose) is removed

• The repressor returns to its original shape and
  binds to the DNA, so that RNA polymerase can no
  longer get past the promoter. No RNA and no
  protein is made.
• Note that RNA polymerase can still bind to the
  promoter though it is unable to move past it.
  That means that when the cell is ready to use the
  operon, RNA polymerase is already there and
  waiting to begin transcription.

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Lac movie
Lac and trp