Multivalency Effects on the Immobilization of Sucrose Phosphorylase in Flow Microchannels and Their Use in the Development of a High-Performance Biocatalytic Microreactor

Donya Valikhani, Juan M. Bolivar, Martin Pfeiffer, Bernd Nidetzky

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Abstract

Microstructured reactors are emerging engineering tools for the development of biocatalytic conversions in continuous flow. A promising layout involves flow microchannels that are wall-coated with enzyme. As protein immobilization within closed microstructures is challenging, we suggested a confluent design of enzyme and microreactor: fusion to the silica-binding module Zbasic2 is used to engineer enzymes for high-affinity oriented attachment to the plain wall surface of glass microchannels. In this study of sucrose phosphorylase, we examined the effects of multiple Zbasic2 modules in a single enzyme molecule on the activity and adsorption stability of the phosphorylase immobilized in a glass microchannel reactor. Compared to the “monovalent” enzyme, two Zbasic2 modules, present in tandem repeat at the N-terminus, separated at the N- and C-terminus of an enzyme monomer, or arranged N-terminally in a protein homodimer, boosted the effectiveness of the immobilized phosphorylase by up to twofold. They attenuated (up to 12-fold) the elution of the wall-coated enzyme during continuous reactor operation. The divalent phosphorylase was distributed uniformly on the microchannel surface and approximately 70 % activity could still be retained after 690 reactor cycles. Reaction–diffusion regime analysis in terms of the second Damköhler number (DaII≤0.02) revealed the absence of mass transport limitations on the conversion rate. The synthesis of α-d-glucose 1-phosphate occurred with a productivity of ∼14 mm min−1 at 50 % substrate conversion (50 mm). The use of wall-coated enzyme microreactors in high-performance biocatalytic reaction engineering is supported strongly.

Original languageEnglish
Pages (from-to)161-166
Number of pages6
JournalChemCatChem
Volume9
Issue number1
DOIs
Publication statusPublished - 9 Jan 2017

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Keywords

  • biocatalysis
  • biotransformations
  • enzymes
  • immobilization
  • microreactors

ASJC Scopus subject areas

  • Catalysis
  • Physical and Theoretical Chemistry
  • Organic Chemistry
  • Inorganic Chemistry

Cooperations

  • NAWI Graz

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