HIGH-YIELD PRODUCTION OF SACCHAROMYCOPSIS-FIBULIGERA GLUCOAMYLASE IN ESCHERICHIA-COLI, REFOLDING, AND COMPARISON OF THE NONGLYCOSYLATED ANDGLYCOSYLATED ENZYME FORMS
A. Solovicova et al., HIGH-YIELD PRODUCTION OF SACCHAROMYCOPSIS-FIBULIGERA GLUCOAMYLASE IN ESCHERICHIA-COLI, REFOLDING, AND COMPARISON OF THE NONGLYCOSYLATED ANDGLYCOSYLATED ENZYME FORMS, Biochemical and biophysical research communications, 224(3), 1996, pp. 790-795
The truncated GLA1 gene encoding the mature form of glucoamylase from
the yeast Saccharomycopsis fibuligera has been over-expressed in Esche
richia coli using the IPTG inducible pET system. Over-expression has l
ed to the accumulation of insoluble glucoamylase in inclusion bodies f
rom which an electrophoretically homogeneous active enzyme has been pr
epared yielding 30 mg per litre medium. This protein represents an N-t
erminus Met-free, non-glycosylated product which displays the identica
l specific activity of 45 units/mg and reduced thermal stability when
compared to glycosylated enzymes isolated from Saccharomyces cerevisia
e carrying the GLA1 gene. These data suggest that S. cerevisiae glycos
ylation of S. fibuligera glucoamylase does not play a critical role in
enzymatic activity but that it does contribute to its thermal stabili
ty. (C) 1996 Academic Press, Inc.