Site-specific mutations of FtsZ--effects on GTPase and in vitro assembly.
BACKGROUND
FtsZ, the major cytoskeletal protein in bacterial cytokinesis, assembles in vitro into protofilaments, which can further associate into sheets, bundles or tubes. We have constructed 16 site-directed mutants of E. coli ftsZ, and tested them for GTP hydrolysis and assembly in vitro, and for their ability to complement the temperature sensitive ftsZ84 mutation in E. coli.


RESULTS
The mutants were grouped into three classes. Benign mutants, which mapped mostly to the front and back surface of the protofilament, were able to complement ftsZ84 in vivo and showed normal assembly in vitro. GTP contact mutations had less than 10% of wild type GTPase activity. They could all assemble in vitro, and several of these mutants could complement ftsZ84. A third, and newly discovered, class of mutations mapped to the sides of the protofilaments. These lateral mutants had mostly normal GTPase and assembly in vitro, but none of them complemented ftsZ84. The non-complementing mutants showed greatly reduced expression from the pBS58 vector, suggesting possible dominant negative effects.


CONCLUSIONS
Several mutants with greatly reduced GTPase could still complement ftsZ84, suggesting that the high level of GTPase observed in vitro is not essential for in vivo function. All of the lateral mutants failed to complement ftsZ84, which suggests that these surfaces of the protofilaments are important for function in cell division. These lateral surfaces may mediate association of FtsZ protofilaments into pairs or small sheets, although their structure is apparently different from the sheets assembled in DEAE dextran or calcium.
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Dept of Cell Biology, 3709 Duke Univ, Med, Cntr, Durham, NC 27710, USA. h.erickson@cellbio.duke.edu
Stricker
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J
Erickson
H P
HP
D002455:Cell Division / Q000502:physiology; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics / Q000378:metabolism*; D020558:GTP Phosphohydrolases / Q000378:metabolism*; D005816:Genetic Complementation Test; D006160:Guanosine Triphosphate / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed
D006160:Guanosine Triphosphate; D020558:GTP Phosphohydrolases