Determination of Apparent Kinetic Parameters and Substrate Specificities of Glucose Isomerase from Streptomyces Roseiscleroticus

Finding suitable sugar replacements, such as syrups, has become necessary due to the rising demand for refined sugar and its high manufacturing costs. Enzymatic technology has been found useful in the production of syrups; one of the useful industrial enzymes for syrup production is glucose isomerase. This study aimed to determine the apparent kinetic parameters and substrate specificities of glucose isomerase from Streptomyces roseiscleroticus. Streptomyces roseiscleroticus with glucose isomerase-producing ability was isolated from soil. The isolated microbial strain was tested for glucose isomerase producing capacity using 2, 3, 5 - triphenyltetrazolium solution as a chromogenic substrate. Crude glucose isomerase was produced, its activity was assayed spectrophotometrically and then partially purified. The apparent kinetic parameters (maximum velocity, Vmax, and Michaelis-Menten constant, KM) of glucose isomerase were determined by nonlinear regression analysis using the Solver add-in function in Microsoft Excel. Estimated Vmax and KM at different concentrations of glucose, xylose, and mannose were 66.621 μmol/min and 0.407 M, 88.478 μmol/min and 0.474 M, 25.016 μmol/min and 1.613 M, respectively. Corresponding substrate specificities 163.554, 186.827 and 15.507 μmolmin-1.M-1 using glucose, xylose, and mannose as substrates, respectively. Comparative evaluation revealed that glucose isomerase from Streptomyces roseiscleroticus has a stronger apparent substrate specificity towards xylose and glucose, and a lower apparent substrate specificity for mannose under the experimental conditions. However, model agreement varied across substrates with glucose providing the most consistent and statistically reliable kinetic characterization.