PURIFICATION AND SPECIFICITY OF RECOMBINANT HORMOCONIS-RESINAE GLUCOAMYLASE-P AND ENDOGENOUS GLUCOAMYLASE FROM TRICHODERMA-REESEI

Hormoconis resinae glucoamylase P of high debranching activity was purified from a recombinant Trichoderma reesei strain. Four different purified fractions were obtained. Three had the same amino terminal sequence as the wild-type enzyme and about the same specific activity, and yielded the same single band on SDS-PAGE after deglycosylation. Presumably they resulted from different glycosylation patterns of the recombinant glucoamylase P. One fraction had a much lower specific activity and yielded tryptic peptides that identified if as the host cellobiohydrolase I contaminated with glucoamylase P. The different glycosylation patterns of recombinant glucoamylase P had only minor effects on its thermal inactivation. During purification of the recombinant glucoamylase, a protein with lower debranching activity was found and purified by chromatofocusing to homogeneity as assessed by SDS-PAGE. It had a pI of about 4.0 and a ratio of pullulan- to starch-degrading activity of 15%. Its amino terminal sequence showed 60% identify to the amino terminal sequence of glucoamylases P and S from Hormoconis resinae. Presumably this enzyme is the endogenous glucoamylase of Trichoderma reesei.