Abstract
To gain further insight into the structural relatedness of tubulin and FtsZ, the tubulin-like prokaryotic cell division protein, we tested the effect of tubulin assembly inhibitors on FtsZ assembly. Common tubulin inhibitors, such as colchicine, colcemid, benomyl, and vinblastine, had no effect on Ca2+-promoted GTP-dependent assembly of FtsZ into polymers. However, the hydrophobic probe 5, 5'-bis-(8-anilino-1-naphthalenesulfonate) (bis-ANS) inhibited FtsZ assembly. The potential mechanisms for inhibition are discussed. Titrations of FtsZ with bis-ANS indicated that FtsZ has one high affinity binding site and multiple low affinity binding sites. ANS (8-anilino-1-naphthalenesulfonate), a hydrophobic probe similar to bis-ANS, had no inhibitory effect on FtsZ assembly. Because tubulin assembly has also been shown to be inhibited by bis-ANS but not by ANS, it supports the idea that FtsZ and tubulin share similar conformational properties. Ca2+, which promotes GTP-dependent FtsZ assembly, stimulated binding of bis-ANS or ANS to FtsZ, suggesting that Ca2+ binding induces changes in the hydrophobic conformation of the protein. Interestingly, depletion of bound Ca2+ with EGTA further enhanced bis-ANS fluorescence. These findings suggest that both binding and dissociation of Ca2+ are capable of inducing FtsZ conformational changes, and these changes could promote the GTP-dependent assembly of FtsZ.
Highlights
Because many inhibitors of tubulin assembly exist, investigating the effects of tubulin inhibitors on FtsZ assembly should provide important information for FtsZ structure and function
Colcemid, benomyl, and vinblastine have no effect on Ca2ϩ-induced FtsZ polymerization, suggesting that FtsZ interacts with these drugs in a manner distinct from that of tubulin
Inhibition of FtsZ Polymerization by Bis-ANS—To study the structural and functional similarity between FtsZ and tubulin, we assessed the effect of tubulin inhibitors on FtsZ polymerization
Summary
Because many inhibitors of tubulin assembly exist, investigating the effects of tubulin inhibitors on FtsZ assembly should provide important information for FtsZ structure and function. We previously developed a simple in vitro assay for FtsZ assembly using a FtsZ-green fluorescent protein fusion (FtsZGFP) [18]. By using this assay, we demonstrated that FtsZ is capable of microtubule-like dynamic assembly and can selfassemble into structures that are similar to microtubule asters. We demonstrated that FtsZ is capable of microtubule-like dynamic assembly and can selfassemble into structures that are similar to microtubule asters This assembly is strictly dependent on GTP. Colcemid, benomyl, and vinblastine have no effect on Ca2ϩ-induced FtsZ polymerization, suggesting that FtsZ interacts with these drugs in a manner distinct from that of tubulin. Because bis-ANS is a fluorescent probe that measures protein hydrophobic surface properties, we used bis-ANS and a related compound, ANS, to probe directly for FtsZ conformational changes induced by Ca2ϩ binding and dissociation
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
