Haloarchaeal-type ?-ketothiolases involved in Poly(3-hydroxybutyrate-co-3-hydroxyvalerate) synthesis in Haloferax mediterranei.

ABSTRACT: The key enzymes for poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) biosynthesis in haloarchaea have been identified except the ?-ketothiolase(s), which condense two acetyl coenzyme A (acetyl-CoA) molecules to acetoacetyl-CoA, or one acetyl-CoA and one propionyl-CoA to 3-ketovaleryl-CoA. Whole-genome analysis has revealed eight potential ?-ketothiolase genes in the haloarchaeon Haloferax mediterranei, among which the PHBV-specific BktB and PhaA were identified by gene knockout and complementation analysis. Unlike all known bacterial counterparts encoded by a single gene, the haloarchaeal PhaA that was involved in acetoacetyl-CoA generation, was composed of two different types of subunits (PhaA? and PhaA?) and encoded by the cotranscribed HFX_1023 (phaA?) and HFX_1022 (phaA?) genes. Similarly, the BktB that was involved in generation of acetoacetyl-CoA and 3-ketovaleryl-CoA, was also composed of two different types of subunits (BktB? and BktB?) and encoded by cotranscribed HFX_6004 (bktB?) and HFX_6003 (bktB?). BktB? and PhaA? were the catalytic subunits and determined substrate specificities of BktB and PhaA, respectively. Their catalytic triad "Ser-His-His" was distinct from the bacterial "Cys-His-Cys." BktB? and PhaA? both contained an oligosaccharide-binding fold domain, which was essential for the ?-ketothiolase activity. Interestingly, BktB? and PhaA? were functionally interchangeable, although PhaA? preferred functioning with PhaA?. In addition, BktB showed biotechnological potential for the production of PHBV with the desired 3-hydroxyvalerate fraction in haloarchaea. This is the first report of the haloarchaeal type of PHBV-specific ?-ketothiolases, which are distinct from their bacterial counterparts in both subunit composition and catalytic residues.

SUBMITTER: Hou J 

PROVIDER: S-EPMC3753943 | biostudies-literature | 2013 Sep

REPOSITORIES: biostudies-literature