Journal of Microbiology

Purification and Properties of Extracellular Poly(3-hydroxybutyrate) Depolymerase Produced by Penicillium pinophilum: Han, Jeen Sun , Son, Young Jong , Chang, Chung Soon , Kim, Mal Nam; J. Microbiol. 1998;36(2):67-73.

Abstract: The extracellular poly(3-hydroxybutyrate)(PHB) depolymerase of Penicilliyum pinophilum ATCC 9644 was purified and characterized. When Penicillum sp. was grown in basal salt medium with PHB as a sole carbon source, higher temperature favored fungal mycelial growth (37℃>30℃>25℃), but enzyme production was lower ar 37℃ than at any other temperatures. The PHB depolymerase was purified using Sepharose CL06B and Sephacryl S0100HR column chromatography. The isolated enzyme was found to be composed of a single polypeptide chain with a molecular weight of about 35 kDa. The optimum condition for the enzyme was pH 6.0 and 50℃, Enzyme activity decreased sharply at temperatures above 50℃. The enzyme was found to be stable in the pH range of 2.0~10.0.1mM Fe^2+ reduced the enzyme activity by 55% abd 4 mM Fe^2+ inhibited it almost completely. The PHB depolymerase (10U) degraded 47% of solvent cast PHB film, while commercial lipase (1000U) of Rhizopus arrhizus degraded 10% of the same specimen, over a period of 48 hours.

Purification and Characterization of Extracellular Poly(3-hydroxybutyrate) Depolymerase from Penicillium simplicissimum LAR13: Jee-Sun Han , Mal-Nam Kim; J. Microbiol. 2002;40(1):20-25.

Abstract: An extracellular PHB depolymerase was purified from P. simplicissimum LAR13 cultural medium by Sepharose CL-6B chromatography. When the fungus was grown in a basal salt medium with poly(3-hydroxybutyrate) (PHB) as the sole carbon source, PHB depolymerase production reached maximum at its stationary phase. The mycelial growth rate was higher at 37 C than at 30 C and even higher than at 25 C, However, the enzyme production was lower at 37 C than 30 C or 25 C. The isolated enzyme is composed of a single polypeptide chain with a molecular mass of about 36 kDa as determined by SDS-PAGE. The optimum conditions for the enzyme activity are pH 5.0 and 45 C. The enzyme was stable for 30 min at a temperature lower than 50 C, and stable at pH higher than 2.0 but it was unstable at pH 1.0. 1 mM Fe^2+ reduced the enzyme activity by 56% and the enzyme was inhibited almost completely by 4 mM Fe^2+ . The enzyme was partially inhibited by phenylmethylsulfonyl fluoride and was very sensitive to diazo-DL-norleucine methyl ester, dithiothreitol and mercuric ion. However, N-p-tosyl-L-lysinechloromethyl ketone, p-hydroxymercuricbenzoate and N-acetylimidazole had no influence upon its activity.

Purification and Characterization of Poly(3-hydroxybutyrate) Depolymerase from a Fungal Isolate, Emericellopsis minima W2: Do Young Kim , Ji Hye Yun , Hyung Woo Kim , Kyung Sook Bae , Young Ha Rhee; J. Microbiol. 2002;40(2):129-133.

Abstract: The fungus, Emericellopsis minima W2, capable of degrading poly(3-hydroxybutyrate) (PHB) was isolated from a waste water sample. Production of the PHB depolymerase from E. minima W2 (PhaZ_Emi ) was significantly repressed in the presence of glucose. PhaZ_Emi was purified by column chromatography on Octyl-Sepharose CL-4B and Sephadex G-100. The molecular mass of the PhaZ_Emi , which consisted of a single polypeptide chain, was estimated to be 48.0 kDa by SDS-PAGE and its pI value was 4.4. The maximum activity of the PhaZ_Emi was observed at pH 9.0 and 55 C. It was significantly inactivated by 1 mM dithiothreitol, 2 mM diisopropyl fluorophosphate, 0.1 mM Tween 80, and 0.1 mM Triton X-100, but insensitive to phenylmethylsulfonyl fluoride and N-ethylmaleimide. The PhaZ_Emi efficiently hydrolyzed PHB and its copolyester with 30 mol% 3-hydroxyvalerate, but did not act on poly(3-hydroxyoctanoate). It also hydrolyzed p-nitrophenylacetate and p-nitrophenylbutyrate but hardly affected the longer-chain forms. The main hydrolysis product of PHB was identified as a dimer of 3-hydroxybutyrate.

Search