IIB   20738
INSTITUTO DE INVESTIGACIONES BIOLOGICAS
Unidad Ejecutora - UE
congresos y reuniones científicas
Título:
Specific contacts between chemoreceptors and the coupling protein CheW are differentially affected during signaling
Autor/es:
ANDREA PEDETTA; CRISTINA CHIAPPE; CLAUDIA STUDDERT
Lugar:
Rosario
Reunión:
Congreso; XIX Congreso Argentino de Microbiología General; 2013
Institución organizadora:
Sociedad Argentina de Microbiología General
Resumen:
Bacterial chemotaxis allows microorganisms to sense
their immediate chemical environment and modulate their movement accordingly.
The main components of this signal transduction system are chemoreceptors,
which sense the chemical stimuli and control the activity of the histidine
kinase CheA, within a ternary complex that also contains the adaptor protein
CheW. The activity of the kinase controls the levels of phosphorylated CheY,
which communicates the sensory input to the flagellar motors.
To accomplish its function, CheW directly interacts
with receptors and with CheA. The domain of CheA that interacts with CheW (P5
domain) is homologous to CheW itself, and both domains show significant
structure and sequence conservation. The interaction of CheW with
chemoreceptors occurs through a hydrophobic surface between the two b-barrels of the protein. CheA is
proposed to interact with the receptors through the corresponding region of its
P5 domain. The actual changes that occur within the ternary complex to modulate
kinase activity during signaling are still being investigated. Several models have
been proposed from structural and biochemical studies using in vitro approaches.
In order to test whether changes in the organization
of the ternary complex can be detected in whole cells, we sought for specific
contacts between chemoreceptors and CheW that could be targets of disulfide
crosslinking. We found pairs of cysteine replacements that form disulfides upon
treatment of whole cells with the oxidant diamide. Disulfide formation was
significantly reduced when a mutant version with altered affinity with
receptors was used, indicating that crosslinking indeed reflected a specific
interaction between both proteins.
Then, we characterized the efficiency of crosslinking
under different conditions. Interestingly, two different crosslinking pairs
showed opposite behavior upon attractant stimuli. This suggests the occurrence
of a signal-dependent relative movement of CheW with respect to the receptors
rather than a change in the association between the two proteins. The
presence/absence of CheA in the cell also affected in opposite ways the
crosslinking efficiency of these two pairs, suggesting that the P5 domain might
compete with CheW more efficiently for one of the interacting points, while
favoring the interaction through the other contact.
The obtained results are discussed under the light of
current experimental evidence.