The gene encoding glucose oxidase (GOD) from Aspergillus niger as expressed as a secretory product in the yeast Saccharomyces cerevisiae. Six consecutive histidine residues were fused to the C-terminus of GOD to facilitate purification, The recombinant GOD-His(6) secreted by cerevisiae migrated as a broad diffuse band on SDS PAGE, with an apparent molecular weight higher than that in natural A. niger GOD. To investigate the effects of hyperglycosylation on the,secretion efficiency and enzyme properties, GOD-His(6) was expressed and secreted in a S. cerevisiae mutant in which the PMR1 gene encoding Ca -ATPase as disrupted. The pmrl null mutant strain secreted an amount of GOD-HiS(6) per unit cell mass higher than that in the wild-type strain. In contrast to the hyperglycosylated GOD-His(6) secreted in the wild-type strain, the pmrl mutant strain secreted GOD-his(6) in a homogeneous form with a protein band pattern similar to that in natural A. niger GOD, based on SDS PAGE. The hyperglycosylated and pmr1Delta mutant-derived GOD-HiS6 enzymes were purified to homogeneity by immobilized metal ion-affinity chromatography and their specific activities and stabilities were compared. The specific activity of the pmr1Delta mutant-derived GOD-His(6) on a protein basis was very similar to that of the hyperglycosylated GOD-HiS(6), although its pH and thermal stabilities were lower than those of the hyperglycosylated GOD-HiS(6). (C) 2002 Elsevier Science (USA). All rights reserved.