A novel glucose oxidase (GOX), a flavoenzyme, from Penicillium sp. was isolated, purified and partially characterised. Maximum activities of 1.08 U mg(-1) dry weight intracellular and 6.9 U ml(-1) extracellular GOX were obtained. Isoelectric focussing revealed two iso-enzymes present in both intra- and extracellular fractions, having pI's of 4.30 and 4.67. GOX from Penicillium sp. was shown to be dimeric with a molecular weight of 148 kDa, consisting of two equal subunits with molecular weight of 70 kDa. The enzyme displayed a temperature optimum between 25 and 30 degrees C, and an optimum pH range of 6-8 for the oxidation of beta-D-glucose. The enzyme was stable at 25 degrees C for a minimum of 10 h, with a half-life of approximately 30 min at 37 degrees C without any prior stabilisation. The lyophilized enzyme was stable at -20 degrees C for a minimum of 6 months. GOX from Penicillium sp. Tt42 displayed the following kinetic characteristics: V., 240.5 U mg(-1); K-m, 18.4 mM; k(cat), 741 s(-1) and k(cat)/K-m, 40 s(-1) mM(-1). Stability at room temperature, good shelf-life without stabilisation and the neutral range for the pH optimum of this GOX contribute to its usefulness in current GOX-based biosensor applications. (c) 2006 Elsevier Inc. All rights reserved.