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, cytochrome c 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 Vs-tyrosinase and phenol. The resulting aggregates can be collected by centrifugation and retain high levels of activity and may find applications in biocatalysis.