Biochemical and functional characterization of the SMC holocomplex from Mycobacterium smegmatis.

Multi-subunit
SMC complexes are required to perform essential functions, such as chromosome compaction, segregation and
DNA repair, from bacteria to humans. Prokaryotic SMC proteins form complexes with two non-SMC
subunits, ScpA and ScpB,
to condense the chromosome. The mutants of both scpa and scpb genes in Bacillus subtilis have been shown to display characteristic
phenotypes such as growth defects and increased frequency of anucleate cells. Here, we studied the function of the
Smc-ScpAB
complex from Mycobacterium smegmatis. We observed no significant growth difference between the scpb null
mutant and wild-type
M. smegmatis under both standard and stress conditions. Furthermore, we characterized the Smc-ScpAB
holocomplex from M. smegmatis. The MsSMC consists of the dimerization hinge and ATPase head domains connected by long
coiled-coils.
The MsSMC interacts with two non-SMC
proteins, ScpA and ScpB, and the resulting holocomplex binds to different
DNA substrates independent of ATP. The Smc-ScpAB
complex showed DNA-stimulated
ATPase activity in the presence of
ssDNA. A cytological profiling assay revealed that upon overexpression the Smc-ScpAB
ternary complex compacts the decondensed
nucleoid of rifampicin-treated
wild-type
and null mukb mutant of Escherichia coli in vivo. Together, our study suggests
that M. smegmatis has a functional Smc-ScpAB
complex capable of DNA binding and condensation. Based on our observations,
we speculate that the presence of alternative SMCs such as MksB or other SMC homologues might have rescued the scpb
mutant phenotype in M. smegmatis.