A Simple Review

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A Simple Review
Tumbling
CM
OM
OM
CM
CW
repellent
ATP
ADP
M
CheY
CheA
CheW
P
MCP
P
RR
Kinase
Smooth Swimming
CM
OM
OM
CM
CCW
attractant
M
CheY
CheA
CheW
MCP
Macnab R. M. Annu. Rev. Microbiol. 2003
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A Simple Review (Cont'd)

• In Salmonella typhimurium, Robert Macnab et al
  found that the temporal gradient apparatus used a
  time-dependent process to sense the change of the
  attractant in its environment.
• Andrew Hughson et al pointed out that the signal
  receiver Trg, in the presence and absence of an
  attractant, had a conformational change mostly
  within a subunit of the homodimer.

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to find out the missing elements in the
apparatusorto study the conformational change
in the MCPs
The next question is
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A ribbon drawing of the ligand-binding domain of
Tar. Shows four alpha helices.
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Dr. Ikuro Kawagishi
Dr. Michael Manson
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Kawagishi's point of view
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• Milligan et al. used artificially
  disulfide-cross-linked dimer of Tar to test for
  its function
• what is the result?
• what can you conclude from the result?

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Their main approach

• Genetics making mutants to study their phenotypes

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Their Tar mutants

• A19K TM1
• T154P
• A198E TM2
• A198E/T154P
• seven deletion derivatives of A198E

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How about Tar-A198E homodimer ?
science 274423-5
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science 274423-5
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Their conclusions

• one of the two cytoplasmic signaling domains in a
  dimer is dispensable for generating an attractant
  response to aspartate
• intersubunit interactions in the "linker" region
  may be required for signaling
• signaling by Tar may be mediated by interaction
  between dimers, conformational changes within a
  single cytoplasmic domain, or both
• a single cytoplasmic domain of a receptor might
  be able to regulate CheA activity at least under
  some conditions

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Manson's point of view
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Evidence against the ligand-induced dimerization
model

• Dimerization states of Tar
• The disulfide-cross-linking experiment
• The heterodimer experiment

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one experiment, two explanations

• oligomerization of Tar cytoplasmic domains
  through leucine zippers strongly stimulates CheA
  in vitro
• what is Manson's explanation ?
• what is Kawagishi's explanation ?

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The same approach

• Genetics

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Their Tar mutants

• R73K
• T154I
• R73K/T154I
• R73K/258oc
• T154I/258oc
• R73K/T154Ioc
• R73K/229oc

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Hydrogen-bonding interaction scheme at the
aspartate binding site. The boxed amino acids are
from subunit A, the ovals, from subunit B. Water
is indicated with W, and hydrogen bonds with
dash lines.
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science 274425-6
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science 274425-6
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Their conclusions

• A heterodimer containing one cytoplasmic domain
  can mediate an attractant signal
• The half-site containing Thr154 may be critical
  for signal propagation and ligand binding
• Region 229-258 is necessary to form functional
  heterodimers
• The cis-inhibition model vs. trans-inhibition
  model

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Common conclusions from two independent research
groups

• one subunit is sufficient for signaling
• T154 is important for both binding and signaling
• the "linker" region is critical for signaling

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• "linker" region the HAMP domain
• Histidine kinases
• Adenylyl cyclases
• Methyl-accepting chemotaxis proteins
• Phosphatases

Aravind 1999
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From piston to rotation
cell 126 829-31
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