Isolation and characterization of polyhydroxyalkanoates inclusions and their associated proteins in Pseudomonas sp. 61-3.
Two types of polyester inclusions of poly(3-hydroxybutyrate) [P(3HB)] and poly(3HB-co-3-hydroxyalkanoates) [P(3HB-co-3HA)] were isolated from crude extract of Pseudomonas sp. 61-3. Proteins associated with each inclusion were separated by SDS-PAGE. PHA synthase 1 (PhaC1(Ps)), PhaF(Ps), and PhaI(Ps) were identified from P(3HB-co-3HA) inclusions by N-terminal amino acid sequences analyses, as well as PHB synthase (PhbC(Ps)) and 24-kDa unknown protein were identified from P(3HB) inclusions. The structural genes of PhaF(Ps) and PhaI(Ps) were located downstream of the pha locus. The relative PHA/PHB synthase activities of each inclusion were measured for various 3-hydroxyacyl-coenzyme As of 4-12 carbon atoms. Direct atomic force microscopy observation of P(3HB) and P(3HB-co-3HA) inclusions demonstrated that the two types of inclusions had different morphologies.
Matsumoto
Ken'ichiro
K
Polymer Chemistry Laboratory, RIKEN Institute, 2-1 Hirosawa, Wako-shi, Saitama 351-0198, Japan.
Matsusaki
Hiromi
H
Taguchi
Kazunori
K
Seki
Minoru
M
Doi
Yoshiharu
Y
D000217:Acyltransferases / Q000302:isolation & purification / Q000378:metabolism; D001426:Bacterial Proteins / Q000302:isolation & purification / Q000378:metabolism; D015153:Blotting, Western; D002479:Inclusion Bodies / Q000737:chemistry* / Q000201:enzymology / Q000648:ultrastructure; D018625:Microscopy, Atomic Force; D011091:Polyesters / Q000138:chemical synthesis / Q000302:isolation & purification*; D011549:Pseudomonas / Q000737:chemistry
D001426:Bacterial Proteins; D011091:Polyesters; D000217:Acyltransferases; C077225:poly(3-hydroxyalkanoic acid) synthase