Novel intracellular 3-hydroxybutyrate-oligomer hydrolase in Wautersia eutropha H16.
Wautersia eutropha H16 (formerly Ralstonia eutropha) mobilizes intracellularly accumulated poly(3-hydroxybutyrate) (PHB) with intracellular poly(3-hydroxybutyrate) depolymerases. In this study, a novel intracellular 3-hydroxybutyrate-oligomer hydrolase (PhaZc) gene was cloned and overexpressed in Escherichia coli. Then PhaZc was purified and characterized. Immunoblot analysis with polyclonal antiserum against PhaZc revealed that most PhaZc is present in the cytosolic fraction and a small amount is present in the poly(3-hydroxybutyrate) inclusion bodies of W. eutropha. PhaZc degraded various 3-hydroxybutyrate oligomers at a high specific activity and artificial amorphous poly(3-hydroxybutyrate) at a lower specific activity. Native PHB granules and semicrystalline PHB were not degraded by PhaZc. A PhaZ deletion mutation enhanced the deposition of PHB in the logarithmic phase in nutrient-rich medium. PhaZc differs from the hydrolases of W. eutropha previously reported and is a novel type of intracellular 3-hydroxybutyrate-oligomer hydrolase, and it participates in the mobilization of PHB along with other hydrolases.
Kobayashi
Teruyuki
T
Laboratory of Molecular Microbiology, Department of Biological Sciences, Faculty of Science, Kanagawa University, 2946 Tsuchiya, Hiratsuka, Kanagawa 259-1293, Japan.
Uchino
Keiichi
K
Abe
Tomoko
T
Yamazaki
Yuya
Y
Saito
Terumi
T
D000595:Amino Acid Sequence; D002265:Carboxylic Ester Hydrolases / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006885:Hydroxybutyrates / Q000378:metabolism*; D002479:Inclusion Bodies / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D010802:Phylogeny; D011091:Polyesters / Q000378:metabolism*; D043367:Ralstonia / Q000201:enzymology / Q000378:metabolism*; D016415:Sequence Alignment; D013379:Substrate Specificity; D000094122:Polyhydroxybutyrates
D002265:Carboxylic Ester Hydrolases; D006885:Hydroxybutyrates; D011091:Polyesters; C003182:poly-beta-hydroxybutyrate; D000094122:Polyhydroxybutyrates; C032152:hydroxybutyrate-dimer hydrolase