Purification and Characterization of an Esterase Involved in Poly(vinyl alcohol) Degradation by Pseudomonas vesicularis PD

An esterase catalyzing the hydrolysis of acetyl ester moieties in poly(vinyl alcohol) was purified 400-fold to electrophoretic homogeneity from the cytoplasmic fraction of Pseudomonas vesicularis PD, which was capable of assimilating poly(vinyl alcohol) as the sole carbon and energy source. The puri...

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Bibliographic Details
Published in:	Bioscience, biotechnology, and biochemistry Vol. 62; no. 10; pp. 2000 - 2007
Main Authors:	SAKAI, Kiyofumi, FUKUBA, Masayuki, HASUI, Yutaka, MORIYOSHI, Kunihiko, OHMOTO, Takashi, FUJITA, Tokio, OHE, Tatsuhiko
Format:	Journal Article
Language:	English
Published:	Tokyo Japan Society for Bioscience, Biotechnology, and Agrochemistry 01-10-1998 Japan Society for Bioscience Biotechnology and Agrochemistry Oxford University Press
Subjects:	Amino Acid Sequence biodegradation Biological and medical sciences Biology of microorganisms of confirmed or potential industrial interest Biotechnology Carboxylic Ester Hydrolases - isolation & purification Carboxylic Ester Hydrolases - metabolism deacetylation esterase Fundamental and applied biological sciences. Psychology Kinetics Mission oriented research Physiology and metabolism poly(vinyl alcohol) Polyvinyl Alcohol - metabolism Pseudomonas - enzymology Pseudomonas vesicularis Substrate Specificity Pseudomonadales Biodegradation Temperature Purification Enzyme Environmental factor Esterases Metabolic pathway Pollutant Enzymatic activity Deacetylation Substrate specificity pH Bacteria Hydrolases Pseudomonadaceae Kinetic parameter Physicochemical properties
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Summary:	An esterase catalyzing the hydrolysis of acetyl ester moieties in poly(vinyl alcohol) was purified 400-fold to electrophoretic homogeneity from the cytoplasmic fraction of Pseudomonas vesicularis PD, which was capable of assimilating poly(vinyl alcohol) as the sole carbon and energy source. The purified enzyme was a homodimeric protein with a molecular mass of 80 kDa and the isoelectric point was 6.8. The pH and temperature optima of the enzyme were 8.0 and 45°C. The enzyme catalyzed the hydrolysis of side chains of poly(vinyl alcohol), short-chain p-nitrophenyl esters, 2-naphthyl acetate, and phenyl acetate, and was slightly active toward aliphatic esters. The enzyme was also active toward the enzymatic degradation products, acetoxy hydroxy fatty acids, of poly(vinyl alcohol). The K m and V max of poly(vinyl alcohol) (degree of polymerization, 500; saponification degree, 86.5-89.0 mol%) and p-nitrophenyl acetate were 0.381% (10.6 mM as acetyl content in the polymer) and 2.56 μM, and 6.52 and 12.6 μmol/min/mg, respectively. The enzyme was strongly inhibited by phenylmethylsulfonyl fluoride and diisopropyl fluorophosphate at a concentration of 5 mM, which indicated that the enzyme was a serine esterase. The pathway for the metabolism of poly(vinyl alcohol) is also discussed.
Bibliography:	ObjectType-Article-2 SourceType-Scholarly Journals-1 ObjectType-Feature-1 content type line 23 ObjectType-Article-1 ObjectType-Feature-2
ISSN:	0916-8451 1347-6947
DOI:	10.1271/bbb.62.2000