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A novel thermostable and efficient Class II glucose isomerase from the thermophilic Caldicoprobacter algeriensis: Biochemical characterization, molecular investigation, and application in High Fructose Syrup production

Author:
Neifar, Sawssan, Hlima, Hajer Ben, Mhiri, Sonia, Mezghani, Monia, Bouacem, Khelifa, Ibrahim, Adel Hadj, Jaouadi, Bassem, Bouanane-Darenfed, Amel, Bejar, Samir
Source:
International journal of biological macromolecules 2019 v.129 pp. 31-40
ISSN:
0141-8130
Subject:
Escherichia coli, active sites, catalytic activity, fructose, genes, glucose, half life, hydrogen bonding, isomerases, molecular models, pH, polypeptides, serine, syrups, thermal stability
Abstract:
A novel glucose isomerase gene from the thermophilic Caldicoprobacter algeriensis, encoding a polypeptide of 438 residues, was identified, cloned and successfully expressed in E. coli. The purified enzyme (GICA) was a homotetramer of about 200 kDa displaying the highest activity at pH 7.0 and 90 °C and retaining 97% of its maximum activity at pH 6.5. The enzyme showed an excellent thermostability with a half-life of 6 min at 100 °C. Interestingly, GICA had a very high affinity of 40 mM and catalytic efficiency of 194 min−1 mM−1 toward d-glucose at 90 °C. A maximum of 54.7% d-glucose to d-fructose conversion was achieved by GICA at 85 °C making it an attractive candidate for HFCS-55 production. The primary sequence inspection and molecular modeling studies revealed that the thermal stability of GICA could be attributed to the presence of extra charged residues at the surface like E108 and Q408 increasing surface charge interactions. Moreover, a serine at position 56 near to P58 could establish hydrogen bond strengthening the dimer attachment. The high catalytic efficiency and affinity of GICA could be ascribed to the presence of amino acid like E108 and K62 that created more charges around the catalytic site entry.
Agid:
6302915