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Purification and kinetics of the PHB depolymerase of Microbacterium paraoxydans RZS6 isolated from a dumping yard

Sayyed, R. Z. and Wani, S. J. and Alyousef, Abdullah A. and Alqasim, Abdulaziz and Syed, Asad and El-Enshasy, Hesham Ali (2019) Purification and kinetics of the PHB depolymerase of Microbacterium paraoxydans RZS6 isolated from a dumping yard. PLoS ONE, 14 (6). e0212324-e0212324. ISSN 1932-6203

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Official URL: http://dx.doi.org/10.1371/journal.pone.0212324

Abstract

Poly-β-hydroxybutyrate (PHB) depolymerase is known to decompose PHB, biodegradable polymers and therefore has great commercial significance in the bioplastic sector. However, reports on PHB depolymerases from isolates obtained from plastic-contaminated sites that reflect the potential of the source organism is scarce. In this study, we evaluated the production of extracellular PHB depolymerase from Microbacterium paraoxydans RZS6 isolated from the plastic-contaminated site in the municipal area of Shahada, Maharashtra, India, for the first time. The isolate was identified using 16S rRNA gene sequencing, gas chromatographic analysis of fatty acid methyl esters (GC-FAME), and BIOLOG method. Ithydro-lyzed PHB on minimal salt medium (MSM) containing PHB as the only source of carbon. The isolate produced PHB depolymerase at 45C during 48 h of incubation. The enzyme was purified most efficiently using octyl-sepharose CL-4B column, with the highest purification yield of 6.675 Umg-1mL-1. The activity of the enzyme was enhanced in the presence of Ca2+ and Mg2+ ions but inhibited by Fe2+ (1 mM) ions and mercaptoethanol (1000 rpm). the nzyme kinetic analysis revealed that the enzyme was a metalloenzyme; requiring Mg2+ ions, that showed optimum enzyme activity at 30C (mesophilic) and under neutrophilic (pH 7) conditions. Scale-up from the shake-flask level to a laboratory-scale bioreactor further enhanced the enzyme yield by 0.809 UmL-1. The molecular weight of the enzyme (40 kDa), as estimated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis, closely resembled the PHB depolymerase of Aureobacterium saperdae. Our findings highlighted the applicability of M. paraoxydans as a producer of extracellular PHB depolymerase having potential of degrading PHB under diverse conditions.

Item Type:Article
Uncontrolled Keywords:PHB depolymerase, Microbacterium paraoxydans
Subjects:T Technology > TP Chemical technology
Divisions:Chemical and Energy Engineering
ID Code:87706
Deposited By: Widya Wahid
Deposited On:30 Nov 2020 21:09
Last Modified:30 Nov 2020 21:09

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