The role of FtsZ-associated proteins in the regulation of the assembly dynamics of Mycobacterium smegmatis FtsZ is not clear. In this work, we examined the effect of M. smegmatis SepF on the assembly and stability of M. smegmatis FtsZ polymers. We discovered a single dominant point mutation in SepF (G51D or G51R) that renders the protein inactive. SepF promoted the polymerization of FtsZ, induced the bundling of FtsZ filaments, stabilized FtsZ filaments and reduced the GTPase activity of FtsZ. Surprisingly, both G51D-SepF and G51R-SepF neither stabilized FtsZ filaments nor showed a discernable effect on the GTPase activity of FtsZ. The binding affinity of SepF to FtsZ was found to be stronger than the binding affinity of G51R/D-SepF to FtsZ. Interestingly, the binding affinity of SepF to G51R-SepF was determined to be 45 times stronger than FtsZ. In addition, the interaction of SepF with G51R-SepF was found to be 2.6 times stronger than SepF–SepF interaction. Furthermore, G51R-SepF impaired the ability of SepF to promote the assembly of FtsZ. In addition, the overexpression of G51R-SepF in M. smegmatis mc2 155 cells retarded the proliferation of these cells and increased the average length of the cells. The results indicated that SepF positively regulates the assembly of M. smegmatis FtsZ and the G51 residue has an important role in the functioning of SepF.
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Cover Image
Cover Image
Cartoon illustration of the simultaneous binding of PAP (a competitive inhibitor of the physiological enzyme co-factor adeonosine 3′-phosphate 5′-phosphosulfate, PAPS) and a heptapeptide saccharide substrate (sticks), poised for sulfation (dashed line) in the active site of heparan sulfate 2-O-sulfotransferase (PDB ID: 4NDZ). The PAP(S)-binding site and the oligosaccharide-binding sites provide dual opportunities for the screening, identification and rational design of small molecule inhibitors of this enzyme, which include the highly sulfated polyanionic drug suramin and the promiscuous protein kinase inhibitor rottlerin. Rottlerin competes with the PAPS co-factor in sulfotransferases, opening the door for the discovery and optimisation of other kinase inhibitors that inhibit these classes of enzyme, including protein tyrosine sulfotransferases (TPSTs). For further details, see the article by Byrne et al. in this issue (pages 2417–2433) and its companion article on pages 2435–2455. Image kindly provided by Neil Berry and Patrick Eyers.
Mutation of G51 in SepF impairs FtsZ assembly promoting ability of SepF and retards the division of Mycobacterium smegmatis cells
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