Immobilization of L-ribose isomerase on the surface of activated mesoporous MCM41 and SBA15 for the synthesis of L-ribose

Abstract

A hexagonal mesoporous molecular sieve-like structure of MCM41 and SBA15 with a large surface area was used to immobilize protein L-ribose isomerase (L-RI) through covalent linkages. The amino group of APTES functionalized nanosilica support MCM41 and SBA15 interacted with glutaraldehyde to promote bidentate linkage and on other side with amino group of enzyme. The use of mesoporous silica matrix for immobilization was observed to conserve the distinctive properties of the protein. The various operational conditions optimized for covalent conjugation of protein with the silica support were found to be dependent on enzyme support ratio, immobilization temperature and time. The immobilization yield of L-RI on MCM41 and SBA15 was achieved to be 60 % (600 mg enzyme /g matrix) and 45 % (450 mg enzyme/g matrix), respectively under the optimized conditions. The immobilized biocatalyst was characterized by various analytical techniques like HR-TEM, EDS, FTIR, TGA and BET. Effects of different experimental conditions were optimized to study enzyme kinetics, pH, temperature, bioconversion, reusability, metal ion effect and storage stability. The biocatalytic efficiency (kcat/Km) was increased by 1.2 fold on immobilization with the catalytic activity of 39.64 IU. Increase in the catalytic efficiency after immobilization could be due to the suitable orientation of enzyme active site and improved accessibility for substrate binding. The immobilization of L-RI on mesoporous silica support could improve the biocatalytic activity, storage stability and reusability. The immobilized biocatalyst was found to be reusable for more than 4 cycles retaining more than 50 % of catalytic activity and promoting the synthesis of a rare sugar L-ribose from L-ribulose with a conversion yield of 22 % in 2 h time.