Activation of glucose isomerase by divalent cations: evidence for two distinct metal-binding sites

Abstract

Activity and metal-content measurements have provided evidence for two distinct metal-binding sites in glucose isomerase from Bacillus coagulans. Although each site binds either Mn 2+ or CO 2+, simultaneous addition of Mn 2+ and Co 2+ to the apoenzyme indicated that one site prefers Mn 2+ (site 1) and the other Co 2+ (site 2). Enzyme activity toward fructose was highest when both sites were filled with Co 2+, whereas the activity toward xylose was highest when site 1 was occupied by Mn 2+. The presence of metal in site 2 did not affect the activity toward xylose. Electron paramagnetic resonance (EPR) spectra of Mn 2+ and Co 2+ tightly bound to the enzyme indicated an asymmetric ligand field, whereas the spectrum of each metal more loosely bound was consistent with a symmetric ligand field. In addition, isomerase activity was reduced by oxidizing Co 2+ to Co 3+ with H 2O 2.