Production and Purification of Poly(3-hydroxybutyrate- co -3-hydroxyvalerate) Degrading Enzyme from Streptomyces sp. AF-
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ORIGINAL PAPER
Production and Purification of Poly(3-hydroxybutyrate-co-3hydroxyvalerate) Degrading Enzyme from Streptomyces sp. AF-111 Siddiq Akbar • Fariha Hasan • Akhtar Nadhman Samiullah Khan • Aamer Ali Shah
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Ó Springer Science+Business Media New York 2013
Abstract A poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV) degrading bacterial strain designated as AF-111 was isolated from sewage sludge sample. The bacterium was identified by 16S rRNA gene sequencing. The results revealed that strain AF-111 showed 99 % similarity with Streptomyces althioticus strain NRRL B-3981 and designated as Streptomyces sp. strain AF-111. An extracellular PHBV depolymerase enzyme was produced under optimized conditions and purified through ammonium sulphate fractionation and column chromatography. The enzyme was purified to homogeneity, indicated by sodium dodecyl sulfate–polyacrylamide gel electrophoresis and molecular weight was found to be approximately 51 kDa. Effect of temperature, pH, metal ions and inhibitors on the PHBV depolymerase activity was determined. The enzyme was stable at wide range of temperature (35–55 °C) and pH (6–8). PHBV depolymerase was stable in the presence of different metal ions except iron and zinc which had inhibitory effect on depolymerase activity. Both ethylenediamine teteracetic acid and phenylmethyl sulphonyl fluoride strongly inhibited enzyme activity which indicates that this enzyme belongs to the serine hydrolase family like other polyhydroxyalkanoate depolymerases. The results show that a depolymerase from strain AF-111 can effectively degrade PHBV, therefore, it can be applied in the process of biochemical monomer recycling. Keywords Poly(3-hydroxybutyrate-co-3hydroxyvalerate) Streptomyces sp. AF-111 PHBV depolymerase SDS-PAGE Metal ions Inhibitors S. Akbar F. Hasan A. Nadhman S. Khan A. A. Shah (&) Department of Microbiology, Faculty of Biological Sciences, Quaid-i-Azam University, Islamabad, 45320, Pakistan e-mail: [email protected]; [email protected]
Introduction Biodegradable polymers are believed to be a suitable, environmentally benign replacement to current petrochemical based polymers [1]. Recent interest in these polymers is provoked on by the increasing costs and eventual depletion of petrochemicals feedstock, concern for the global environment and greenhouse gases, and a general paradigm shift towards sustainable manufacturing [2]. To date, the applications for biodegradable polymers have grown to include mainly agricultural, biomedical, and food packaging [3, 4]. Polyhydroxyalkanoates (PHAs) are natural biodegradable polymers which are synthesized and accumulated intracellularly as a carbon and energy reserve material, during unbalanced growth conditions by a wide variety of bacteria and fungi [5]. PHAs are produced by a number of microorganisms on a variety of substrates, including renewable and fossil resources. Genetically engineered microorganisms and plants are also used for the production of PHAs [6]. PHAs exhibit material properties similar t
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