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bioRxiv · 10.1101/698381

Production, purification and evaluation of biodegrading potential of PHB depolymerase of Stenotrophomonas sp. RZS 7

Sayyed, R. Z.·Wani, S.·Al-Harthi, H.·Syed, A.·El-Enshasy, H. A.
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Abstract

There are numerous reports on PHB depolymerases produced by a wide variety of microorganisms isolated from various habitats, however, reports on PHB depolymerase isolated from plastic contaminated sites are scares. Thermophilic PHB polymerase produced by isolates obtained from plastic contaminated sites is expected to have better relevance for its application in plastic/bioplastic degradation. Although PHB has attracted commercial significance, the inefficient production and recovery methods, inefficient purification of PHB depolymerase and lack of ample knowledge on PHB degradation by PHB depolymerase have hampered its large scale commercialization. Therefore, to ensure the biodegradability of biopolymers, it becomes imperative to study the purification of the biodegrading enzyme system. We report the production, purification, and characterization of extracellular PHB depolymerase from Stenotrophomonas sp. RZS 7 isolated from a plastic contaminated site. The isolate produced extracellular poly-{beta}-hydroxybutyrate (PHB) depolymerase in the mineral salt medium at 30oC during 4 days of incubation under shake flask condition. Purification of the enzyme was carried out by three different methods using PHB as a substrate. Purification of PHB depolymerase by ammonium salt precipitation, column chromatography, and solvent purification method was successfully carried out. Among the purification method tested, the enzyme was best purified by column chromatography on Octyl-Sepharose CL-4B column with maximum (0.7993 U mg-1 ml-1) purification yield. The molecular weight of purified PHB depolymerase (40 kDa) closely resembled with PHB depolymerase of Aureobacterium saperdae. Experiments on assessment of biodegradation of PHB in liquid culture medium and under natural soil conditions confirmed PHB biodegradation potential of Stenotrophomonas sp. RZS 7. The results obtained in FTIR analysis, HPLC study and GC-MS analysis confirmed the biodegradation attempt in liquid medium by Stenotrophomonas sp. RZS 7. Changes in surface morphology of PHB film in soil burial as observed in FE SEM analysis confirmed the biodegradation of PHB. The isolate was capable of degrading PHB and resulted in 87.74% degradation. Higher rate of degradation under natural soil condition is the result of activity of soil microbes that complemented the degradation by Stenotrophomonas sp. RZS 7.

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Sayyed, R. Z., Wani, S., Al-Harthi, H., Syed, A., El-Enshasy, H. A.. 2019-07-10. Production, purification and evaluation of biodegrading potential of PHB depolymerase of Stenotrophomonas sp. RZS 7. https://doi.org/10.1101/698381

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