Targeting the chromosome partitioning protein ParA in tuberculosis drug discovery.
OBJECTIVE
To identify inhibitors of the essential chromosome partitioning protein ParA that are active against Mycobacterium tuberculosis.


METHODS
Antisense expression of the parA orthologue MSMEG_6939 was induced on the Mycobacterium smegmatis background. Screening of synthetic chemical libraries was performed to identify compounds with higher anti-mycobacterial activity in the presence of parA antisense. Differentially active compounds were validated for specific inhibition of purified ParA protein from M. tuberculosis (Rv3918c). ParA inhibitors were then characterized for their activity towards M. tuberculosis in vitro.


RESULTS
Under a number of culture conditions, parA antisense expression in M. smegmatis resulted in reduced growth. This effect on growth provided a basis for the detection of compounds that increased susceptibility to expression of parA antisense. Two compounds identified from library screening, phenoxybenzamine and octoclothepin, also inhibited the in vitro ATPase activity of ParA from M. tuberculosis. Structural in silico analyses predict that phenoxybenzamine and octoclothepin undergo interactions compatible with the active site of ParA. Octoclothepin exhibited significant bacteriostatic activity towards M. tuberculosis.


CONCLUSIONS
Our data support the use of whole-cell differential antisense screens for the discovery of inhibitors of specific anti-tubercular drug targets. Using this approach, we have identified an inhibitor of purified ParA and whole cells of M. tuberculosis.
Nisa
Shahista
S
School of Biological Sciences, Victoria University of Wellington, Wellington, New Zealand.
Blokpoel
Marian C J
MC
Robertson
Brian D
BD
Tyndall
Joel D A
JD
Lun
Shichun
S
Bishai
William R
WR
O'Toole
Ronan
R
D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism; D000995:Antitubercular Agents / Q000494:pharmacology*; D001426:Bacterial Proteins / Q000037:antagonists & inhibitors* / Q000378:metabolism; D020090:Chromosome Segregation / Q000187:drug effects*; D003988:Dibenzothiepins / Q000494:pharmacology; D004353:Drug Evaluation, Preclinical / Q000379:methods*; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D055785:Gene Knockdown Techniques; D006801:Humans; D020102:Mycobacterium smegmatis / Q000187:drug effects* / Q000254:growth & development; D009169:Mycobacterium tuberculosis / Q000187:drug effects* / Q000254:growth & development; D010643:Phenoxybenzamine / Q000494:pharmacology; D016372:RNA, Antisense / Q000096:biosynthesis / Q000235:genetics
D000995:Antitubercular Agents; D001426:Bacterial Proteins; D003988:Dibenzothiepins; D004791:Enzyme Inhibitors; D016372:RNA, Antisense; C052883:chromosome partition proteins, bacterial; D010643:Phenoxybenzamine; C100288:octoclothepine; D000251:Adenosine Triphosphatases
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