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Genetic Analysis of the Escherichia coli FtsZ·ZipA Interaction in the Yeast Two-hybrid System

S. Haney,E. Glasfeld,C. Hale,D. Keeney,Zhizhen He,P. D. de Boer

Published 2001 in Journal of Biological Chemistry

ABSTRACT

The recruitment of ZipA to the septum by FtsZ is an early, essential step in cell division in Escherichia coli. We have used polymerase chain reaction-mediated random mutagenesis in the yeast two-hybrid system to analyze this interaction and have identified residues within a highly conserved sequence at the C terminus of FtsZ as the ZipA binding site. A search for suppressors of a mutation that causes a loss of interaction (ftsZ D373G) identified eight different changes at two residues within this sequence. In vitro, wild type FtsZ interacted with ZipA with a high affinity in an enzyme-linked immunosorbent assay, whereas FtsZD373G failed to interact. Two mutant proteins examined restored this interaction significantly.In vivo, the alleles tested are significantly more toxic than the wild type ftsZ and cannot complement a deletion. We have shown that a fusion, which encodes the last 70 residues of FtsZ in the two-hybrid system, is sufficient for the interaction with FtsA and ZipA. However, when the wild type sequence is compared with one that encodes FtsZD373G, no interaction was seen with either protein. Mutations surrounding Asp-373 differentially affected the interactions of FtsZ with ZipA and FtsA, indicating that these proteins bind the C terminus of FtsZ differently.

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