Changing the direction of flagellar rotation in bacteria by modulating the ratio between the rotational states of the switch protein FliM.
One of the major questions in bacterial chemotaxis is how the switch, which controls the direction of flagellar rotation, functions. It is well established that binding of the signaling molecule CheY to the switch protein FliM shifts the rotation from the default direction, counterclockwise, to clockwise. How this shift is done is still a mystery. Our aim in this study was to determine the correlation between the fraction of FliM molecules in the clockwise state (i.e. occupied by CheY) and the probability of clockwise rotation. For this purpose we gradually expressed, from a plasmid, a clockwise FliM mutant protein in cells that express, from the chromosome, wild-type FliM but no chemotaxis proteins. We verified that plasmid-borne FliM exchanges chromosomal FliM in the switch. Surprisingly, a substantial clockwise probability was not obtained before the large majority of the FliM molecules in the switch were clockwise molecules. Thereafter, the rise in clockwise probability was very steep. These results suggest that an increase in the clockwise probability requires a high level of FliM occupancy by CheY approximately P. They further suggest that the steep increase in clockwise rotation upon increasing CheY levels, reported in several studies, is due, at least in part, to cooperativity of post-binding interactions within the switch. We also carried out the inverse experiment, in which wild-type FliM was gradually expressed in a background of a clockwise fliM mutant. In this case, the level of the clockwise mutant protein, required for establishing a certain clockwise probability, was lower than in the original experiment. If our system (in which the ratio between the rotational states of FliM in the switch is established by slow exchange) and the native system (in which the ratio is established by fast changes in FliM occupancy) are comparable, the results suggest that hysteresis is involved in the switch function. Such a situation might reflect a damping mechanism, which prevents a situation in which fluctuations in the phosphorylation level of CheY throw the switch from one direction of rotation to the other.
Bren
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A
Department of Biological Chemistry, The Weizmann Institute of Science,76100 Rehovot, Israel.
Eisenbach
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M
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004755:Enterobacteriaceae / Q000166:cytology* / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000166:cytology / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins; D005407:Flagella / Q000502:physiology*; D008565:Membrane Proteins / Q000378:metabolism; D000072236:Methyl-Accepting Chemotaxis Proteins; D010766:Phosphorylation; D011336:Probability; D011485:Protein Binding; D012399:Rotation; D012475:Salmonella / Q000166:cytology / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction
D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D008565:Membrane Proteins; D000072236:Methyl-Accepting Chemotaxis Proteins; C000607606:cheY protein, E coli; C066821:FliM protein, Bacteria