Immobilisation and kinetics of monoamine oxidase (MAO-N-D5) enzyme in polyvinyl alcohol gels

Research output: Contribution to journalArticle

  • External authors:
  • Kristína Markošová
  • Igor Dolejš
  • Radek Stloukal
  • Leonardo Rios-Solis
  • Michal Rosenberg
  • Martina Micheletti
  • Gary J. Lye
  • Martin Rebroš

Abstract

This study focused on the production and immobilisation of the crude enzyme extract of recombinant monoamine oxidase (EC 1.4.3.4), originating from Aspergillus Niger (MAO-N-D5) and expressed in Escherichia coli, in PVA gel using the LentiKats® technique. MAO-Ns are important enzymes in the chemical industry due to their stereoselectivity and they are often used for the deracemisation of non-optically pure mixtures of amines. Biomass production, enzyme preparation, enzyme immobilisation, process parameters for the immobilised enzyme and characterisation of the enzyme are described in detail here. The biomass was prepared in laboratory bioreactors, and two different disruption techniques were compared. The activity of the enzyme was determined by biotransformation with secondary amine 3-azabicyclo [3,3,0] octane as a substrate. The crude enzyme extract showed 61.5% of the whole cell activity and the immobilised enzyme showed a wider optimum pH and temperature ranges than the free enzyme. The initial specific activity of the immobilised monoamine oxidase crude enzyme extract remained at 80% after 12 repeated biotransformations. For the first time, the full kinetic parameters of an immobilised MAO-N-D5 were obtained based on a ping-pong bi-bi reaction mechanism. The specific activity was 0.29 U g-1 (Lentikats) and the Km was 7.31 mM, which were similar in comparison to whole cell MAO-N-D5. Characterisation of immobilised MAO-N-D5 showed particular benefits in terms of activity and stability in comparison with free and whole cell MAO-N-D5; therefore, the immobilisation of this enzyme is very suitable for industrial applications.

Bibliographical metadata

Original languageEnglish
Pages (from-to)69-74
Number of pages6
JournalJournal of Molecular Catalysis - B Enzymatic
Volume129
Early online date19 Apr 2016
DOIs
StatePublished - 1 Jul 2016