A Novel Affinity Chromatographic Material for the Purification of Extracellular Polyhydroxybutyrate Depolymerases

Christos P. Papaneophytou, Anastasia A. Pantazaki

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3 Citations (Scopus)

Abstract

A novel affinity chromatographic material, which is composed of silica matrix, coated with polyhydroxybutyrate (PHB) powder, suitable for the purification of PHB depolymerases, was developed. The surface morphology of the PHB-silica coated particles (silica-PHB composite particles) was examined by scanning electron microscopy and revealed a successful uniform coating of silica particles with PHB. Moreover, the complex of these materials retained its homogeneity even after incubation at 80 °C for 6 h, whereas the strong binding of PHB on silica surface was further verified by thermal gravimetric analysis and by PHB extraction- from silica surface- experiments. This novel material was demonstrated to be suitable for both, the one-step on-batch and on-column purification of Thermus thermophilus extracellular PHB depolymerase. The enzyme exhibited higher affinity against the composite of silica-PHB particles than PHB powder, since the one-step purification-fold and the overall recovery of the enzyme were 2.8 and 4 times higher respectively, in the first case. Reusability of the silica-PHB composites particles was examined by determining the recoveries of PHB depolymerase. The enzyme recoveries were ranged from 30 to 35% for the first five uses, whereas for further uses recoveries gradually dropped to 15-18% indicating that the particles could be used repeatedly for five times. This material could be also a suitable support for lipases or other proteins that exhibit strong affinity to hydrophobic materials.

Original languageEnglish
Pages (from-to)876-886
Number of pages11
JournalJournal of Polymers and the Environment
Volume19
Issue number4
DOIs
Publication statusPublished - Dec 2011

Keywords

  • Affinity chromatography
  • Extracellular PHB-depolymerase
  • Silica-PHB composite particles
  • Thermus thermophilus HB8

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