Recombinant Hansenula polymorpha as a biocatalyst: coexpression of the spinach glycolate oxidase ( GO ) and the S. cerevisiae catalase T ( CTT1 ) gene

Abstract

The methylotrophic yeast Hansenula polymorpha has been developed as an efficient production system for heterologous proteins. The system offers the possibility to cointegrate heterologous genes in anticipated fixed copy numbers into the chromosome. As a consequence co-production of different proteins in stoichiometric ratios can be envisaged. This provides options to design this yeast as an industrial biocatalyst in procedures where several enzymes are required for the efficient conversion of a given inexpensive compound into a valuable product. To this end recombinant strains have been engineered with multiple copies of expression cassettes containing the glycolate oxidase (GO) gene from spinach and the catalase T (CTT1) gene from S. cerevisiae. The newly created strains produce high levels of the peroxisomal glycolate oxidase and the cytosolic catalase T. The strains efficiently convert glycolate into glyoxylic acid, oxidizing the added substrate and decomposing the peroxide formed during this reaction into water and oxygen.

The methylotrophic yeast Hansenula polymorpha: a versatile cell factory

“…a novel Hepatitis B vaccine) [52,53] and others will soon follow. Also, the use of H. polymorpha as a biocatalyst [54] will be extended in the near future in particular when the option to anchor factors involved in the activation of complex heterologous proteins at the periplasmic space or outer cell wall will be realized. This strategy allows the creation of regeneratable systems in continuous cultures.…”

Uricase production by a recombinant Hansenula polymorpha strain harboring Candida utilis uricase gene

“…As the methylotrophic yeast, H. polymorpha in particular has been studied extensively for its highly regulated biogenesis of peroxisomes, nitrate assimilation, and methanol metabolism. The expression of the key methanolmetabolizing enzymes, such as methanol oxidase (MOX; Ledeboer et al 1985) and formate dehydrogenase (Gellissen et al 1996), can amount up to one third of the total cellular protein during growth on methanol. The existence of inducible strong promoters, the formation of integrants in high copy numbers, high cell density (>100 g dry weight per liter) in cheap defined media, and the low antigenicity of proteins produced in H. polymorpha make H. polymorpha a highly competitive system for the production of recombinant proteins for medical application (Gellissen 2000;Gellissen et al 2005).…”

Hydrogen Peroxide in Biocatalysis. A Dangerous Liaison

“…A recombinant Hansenula polymorpha expressing the spinach glycolate oxidase and the Saccharomyces cerevisiae catalase T genes were used to convert glycolate into glyoxylic acid [182]. Trigonopsis variabilis induced for D-amino acid oxidase and catalase were immobilized by entrapment in polyacrylamide beads obtained by radiation polymerization [183].…”