Cysteine 155 plays an important role in the assembly of Mycobacterium tuberculosis FtsZ

Abstract

The assembly of FtsZ plays an important role in bacterial cell division. Mycobacterium tuberculosis FtsZ ( Mtb FtsZ) has a single cysteine residue at position 155. We have investigated the role of the lone cysteine residue in the assembly of Mtb FtsZ using different complimentary approaches, namely chemical modification by a thiol‐specific reagent 5,5′‐dithiobis‐(2‐nitrobenzoic acid) (DTNB) or a cysteine‐chelating agent HgCl 2 , and site‐directed mutagenesis of the cysteine residue. HgCl 2 strongly reduced the polymerized mass of Mtb FtsZ while it had no detectable effect on the polymerization of Escherichia coli FtsZ, which lacks a cysteine residue. HgCl 2 inhibited the protofilamentous assembly of Mtb FtsZ and induced the aggregation of the protein. Further, HgCl 2 perturbed the secondary structure of Mtb FtsZ and increased the binding of a hydrophobic probe 1‐anilinonaphthalene‐8‐sulfonic acid (ANS) with Mtb FtsZ, indicating that the binding of HgCl 2 altered the conformation of Mtb FtsZ. Chemical modification of Mtb FtsZ by DTNB also decreased the polymerized mass of Mtb FtsZ. Further, the mutagenesis of Cys‐155 to alanine caused a strong reduction in the assembly of Mtb FtsZ. Under assembly conditions, the mutated protein formed aggregates instead of protofilaments. Far‐UV CD spectroscopy and ANS binding suggested that the mutated Mtb FtsZ has different conformation than that of the native Mtb FtsZ. The effect of the mutation or chemical modification of Cys‐155 on the Mtb FtsZ assembly has been explained considering its location in the Mtb FtsZ crystal structure. The results together suggest that the cysteine residue (Cys‐155) of Mtb FtsZ plays an important role in the assembly of Mtb FtsZ into protofilaments.