Characterization of the highly active polyhydroxyalkanoate synthase of Chromobacterium sp. strain USM2.

TitleCharacterization of the highly active polyhydroxyalkanoate synthase of Chromobacterium sp. strain USM2.
Publication TypeJournal Article
Year of Publication2011
AuthorsBhubalan, K, Chuah, J-A, Shozui, F, Brigham, CJ, Taguchi, S, Sinskey, AJ, Rha, CK, Sudesh, K
JournalAppl Environ Microbiol
Date Published2011 May
Keywords3-Hydroxybutyric Acid, Acyltransferases, Caproates, Chromobacterium, Cloning, Molecular, DNA, Bacterial, Escherichia coli, Kinetics, Molecular Sequence Data, Pentanoic Acids, Recombinant Proteins, Sequence Analysis, DNA, Substrate Specificity

The synthesis of bacterial polyhydroxyalkanoates (PHA) is very much dependent on the expression and activity of a key enzyme, PHA synthase (PhaC). Many efforts are being pursued to enhance the activity and broaden the substrate specificity of PhaC. Here, we report the identification of a highly active wild-type PhaC belonging to the recently isolated Chromobacterium sp. USM2 (PhaC(Cs)). PhaC(Cs) showed the ability to utilize 3-hydroxybutyrate (3HB), 3-hydroxyvalerate (3HV), and 3-hydroxyhexanoate (3HHx) monomers in PHA biosynthesis. An in vitro assay of recombinant PhaC(Cs) expressed in Escherichia coli showed that its polymerization of 3-hydroxybutyryl-coenzyme A activity was nearly 8-fold higher (2,462 ± 80 U/g) than that of the synthase from the model strain C. necator (307 ± 24 U/g). Specific activity using a Strep2-tagged, purified PhaC(Cs) was 238 ± 98 U/mg, almost 5-fold higher than findings of previous studies using purified PhaC from C. necator. Efficient poly(3-hydroxybutyrate) [P(3HB)] accumulation in Escherichia coli expressing PhaC(Cs) of up to 76 ± 2 weight percent was observed within 24 h of cultivation. To date, this is the highest activity reported for a purified PHA synthase. PhaC(Cs) is a naturally occurring, highly active PHA synthase with superior polymerizing ability.

Alternate JournalAppl Environ Microbiol
Citation Key111
PubMed ID21398494
PubMed Central IDPMC3126384