1887

Abstract

Branched-chain amino acids (BCAAs) are essential amino acids, but their biosynthetic pathway is absent in mammals. Ketol-acid reductoisomerase (IlvC) is a BCAA biosynthetic enzyme that is coded by Rv3001c in H37Rv (Rv) and MRA_3031 in H37Ra (Ra). IlvCs are essential in Rv as well as in . Compared to wild-type and IlvC-complemented -Ra strains, IlvC knockdown strain showed reduced survival at low pH and under low pH+starvation stress conditions. Further, increased expression of IlvC was observed under low pH and starvation stress conditions. Confirmation of a role for IlvC in pH and starvation stress was achieved by developing BL21(DE3) IlvC knockout, which was defective for growth in M9 minimal medium, but growth could be rescued by isoleucine and valine supplementation. Growth was also restored by complementing with over-expressing constructs of Ra and IlvCs. The knockout also had a survival deficit at pH=5.5 and 4.5 and was more susceptible to killing at pH=3.0. The biochemical characterization of Ra and IlvCs confirmed that both have NADPH-dependent activity. In conclusion, this study demonstrates the functional complementation of IlvC by Ra IlvC and also suggests that IlvC has a role in tolerance to low pH and starvation stress.

Funding
This study was supported by the:
  • Science and Engineering Research Board (Award EMR/20l7/001295)
    • Principle Award Recipient: Kumar SinghSudheer
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2021-09-13
2024-03-29
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