Journal
JOURNAL OF BIOLOGICAL CHEMISTRY
Volume 282, Issue 38, Pages 27736-27743Publisher
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
DOI: 10.1074/jbc.M703788200
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Funding
- NIAID NIH HHS [R01 AI048417-06A1, R01 AI048417, R21 AI064502, AI64502, AI48417] Funding Source: Medline
- NIGMS NIH HHS [GM66014, R01 GM066014, R01 GM066014-04] Funding Source: Medline
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We have investigated the assembly of FtsZ from Mycobacterium tuberculosis (MtbFtsZ). Electron microscopy confirmed the previous observation that MtbFtsZ assembled into long, two-stranded filaments at PH 6.5. However, we found that assembly at pH 7.2 or 7.7 produced predominantly short, one-stranded protofilaments, similar to those of Escherichia coli FtsZ (EcFtsZ). Near pH 7, which is close to the pH of M. tuberculosis cytoplasm, MtbFtsZ formed a mixture of single- and two-stranded filaments. We developed a fluorescence resonance energy transfer assay to measure the kinetics of initial assembly and the dynamic properties at steady state. Assembly of MtbFtsZ reached a plateau after 60-100 s, about 10 times slower than EcFtsZ. The initial assembly kinetics were similar at pH 6.5 and 7.7, despite the striking difference in the polymer structures. Both were fit with a cooperative assembly mechanism involving a weak dimer nucleus, similar to EcFtsZ but with slower kinetics. Subunit turnover and GTPase at steady state were also about 10 times slower for MtbFtsZ than for EcFtsZ. Specifically, the half-time for subunit turnover in vitro at PH 7.7 was 42 s for MtbFtsZ compared with 5.5 s for EcFtsZ. Photo-bleaching studies in vivo showed a range of turnover half-times with an average of 25 s for MtbFtsZ as compared with 9 s for EcFtsZ.
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