Production of chiral (R)-3-hydroxyoctanoic acid monomers, catalyzed by Pseudomonas fluorescens GK13 poly(3-hydroxyoctanoic acid) depolymerase.
The extracellular medium-chain-length polyhydroxyalkanoate (MCL-PHA) depolymerase of Pseudomonas fluorescens GK13 catalyzes the hydrolysis of poly(3-hydroxyoctanoic acid) [P(3HO)]. Based on the strong tendency of the enzyme to interact with hydrophobic materials, a low-cost method which allows the rapid and easy purification and immobilization of the enzyme has been developed. Thus, the extracellular P(3HO) depolymerase present in the culture broth of cells of P. fluorescens GK13 grown on mineral medium supplemented with P(3HO) as the sole carbon and energy source has been tightly adsorbed onto a commercially available polypropylene support (Accurel MP-1000) with high yield and specificity. The activity of the pure enzyme was enhanced by the presence of detergents and organic solvents, and it was retained after treatment with an SDS-denaturing cocktail under both reducing and nonreducing conditions. The time course of the P(3HO) hydrolysis catalyzed by the soluble and immobilized enzyme has been assessed, and the resulting products have been identified. After 24 h of hydrolysis, the dimeric ester of 3-HO [(R)-3-HO-HO] was obtained as the main product of the soluble enzyme. However, the immobilized enzyme catalyzes almost the complete hydrolysis of P(3HO) polymer to (R)-3-HO monomers under the same conditions.
Gangoiti
Joana
J
Enzyme and Cell Technology Group, Department of Biochemistry and Molecular Biology, Faculty of Science and Technology, University of the Basque Country, Bilbao, Spain.
Santos
Marta
M
Llama
María J
MJ
Serra
Juan L
JL
D001426:Bacterial Proteins / Q000302:isolation & purification / Q000378:metabolism*; D002210:Caprylates / Q000378:metabolism*; D002265:Carboxylic Ester Hydrolases / Q000302:isolation & purification / Q000378:metabolism*; D003470:Culture Media / Q000737:chemistry; D004800:Enzymes, Immobilized / Q000302:isolation & purification / Q000378:metabolism; D011551:Pseudomonas fluorescens / Q000201:enzymology*; D013997:Time Factors
D001426:Bacterial Proteins; D002210:Caprylates; D003470:Culture Media; D004800:Enzymes, Immobilized; C101940:3-hydroxyoctanoic acid; D002265:Carboxylic Ester Hydrolases; C080782:poly(3-hydroxyoctanoic acid) depolymerase
Antonie Van Leeuwenhoek. 2003;83(2):183-9
12785312
Enzyme Microb Technol. 1996 May 1;18(6):392-416
8936427
Chembiochem. 2002 Jun 3;3(6):566-71
12325013
J Microbiol. 2007 Apr;45(2):87-97
17483792
J Appl Microbiol. 2007 Jun;102(6):1437-49
17578408
Biotechnol Lett. 2006 Mar;28(6):383-8
16614903
Annu Rev Microbiol. 2002;56:403-32
12213937
Biomacromolecules. 2002 Mar-Apr;3(2):291-6
11888314
Appl Environ Microbiol. 1995 Aug;61(8):3113-8
7487042
J Biotechnol. 1998 Oct 27;65(2-3):127-61
9828458
Arch Microbiol. 2004 Oct;182(2-3):212-25
15340783
Appl Microbiol Biotechnol. 2000 Mar;53(3):323-7
10772473
Chem Soc Rev. 2009 Aug;38(8):2434-46
19623359
J Microbiol. 2005 Jun;43(3):285-94
15995648
Biotechnol Bioeng. 2002 Mar 20;77(6):717-22
11807767
Microbiol Rev. 1990 Dec;54(4):450-72
2087222
J Bacteriol. 1994 Nov;176(22):7065-73
7961472
Nature. 1970 Aug 15;227(5259):680-5
5432063
Methods Enzymol. 1983;91:95-119
6855607
Appl Microbiol Biotechnol. 1996 Dec;46(5-6):451-63
9008883
Appl Environ Microbiol. 2006 Sep;72(9):6094-100
16957234
Appl Microbiol Biotechnol. 2005 Jun;67(5):648-53
15711795
Trends Biotechnol. 1998 Oct;16(10):419-27
9807839
J Bacteriol. 1995 Feb;177(3):596-607
7836292
J Biosci Bioeng. 2000;89(2):196-8
16232726
Appl Environ Microbiol. 1993 Apr;59(4):1220-7
8476295
Appl Microbiol Biotechnol. 2005 Jun;67(5):592-9
15700123