Cross-linking and immobilisation of different proteins with recombinant Verrucomicrobium spinosum tyrosinase

This paper reports on the cross-linking and immobilisation of various proteins by the recombinant tyrosinase from Verrucomicrobium spinosum (Vs-tyrosinase). In general it is found that Vs-tyrosinase can readily cross-link proteins with a low degree of complexity, such as casein, but that the enzyme cannot readily cross-link well folded protein substrates such as lysozyme, myoglobin, cytochromec or Candida antarctica lipase B (CALB). However, the inclusion of phenolic compounds (phenol or caffeic acid) to reaction mixtures of these proteins can greatly enhance the levels of cross-linking. For example it is possible to prepare cross-linked aggregates of industrially applicable enzymes such as CALB by simply incubating it with Vstyrosinase and phenol. The resulting aggregates can be collected by centrifugation and retain high levels of activity and may find applications in biocatalysis. (C) 2010 Elsevier B.V. All rights reserved.