title: A highly efficient sorbitol dehydrogenase from Gluconobacter oxydans G624 and improvement of its stability through immobilization.
abstract: A sorbitol dehydrogenase (GoSLDH) from Gluconobacter oxydans G624 (G. oxydans G624) was expressed in Escherichia coli BL21(DE3)-CodonPlus RIL. The complete 1455-bp codon-optimized gene was amplified, expressed, and thoroughly characterized for the first time. GoSLDH exhibited Km and kcat values of 38.9 mM and 3820 s(-1) toward L-sorbitol, respectively. The enzyme exhibited high preference for NADP(+) (vs. only 2.5% relative activity with NAD(+)). GoSLDH sequencing, structure analyses, and biochemical studies, suggested that it belongs to the NADP(+)-dependent polyol-specific long-chain sorbitol dehydrogenase family. GoSLDH is the first fully characterized SLDH to date, and it is distinguished from other L-sorbose-producing enzymes by its high activity and substrate specificity. Isothermal titration calorimetry showed that the protein binds more strongly to D-sorbitol than other L-sorbose-producing enzymes, and substrate docking analysis confirmed a higher turnover rate. The high oxidation potential of GoSLDH for D-sorbitol was confirmed by cyclovoltametric analysis. Further, stability of GoSLDH significantly improved (up to 13.6-fold) after cross-linking of immobilized enzyme on silica nanoparticles and retained 62.8% residual activity after 10 cycles of reuse. Therefore, immobilized GoSLDH may be useful for L-sorbose production from D-sorbitol.
authors.affiliation: Department of Chemical Engineering, Konkuk University, Seoul 05029, Korea.
authors.forename: Tae-Su
authors.initials: TS
authors.lastname: Kim
authors.affiliation: Department of Chemical Engineering, Konkuk University, Seoul 05029, Korea.
authors.forename: Sanjay K S
authors.initials: SK
authors.lastname: Patel
authors.affiliation: Department of Chemical Engineering, Konkuk University, Seoul 05029, Korea.
authors.forename: Chandrabose
authors.initials: C
authors.lastname: Selvaraj
authors.affiliation: Systems Biotechnology Research Center, KIST Gangneung Institute of Natural Products, 25451, Republic of Korea.
authors.forename: Woo-Suk
authors.initials: WS
authors.lastname: Jung
authors.affiliation: Systems Biotechnology Research Center, KIST Gangneung Institute of Natural Products, 25451, Republic of Korea.
authors.forename: Cheol-Ho
authors.initials: CH
authors.lastname: Pan
authors.affiliation: Department of Materials Science and Engineering, Korea University, Seoul 02841, Republic of Korea.
authors.forename: Yun Chan
authors.initials: YC
authors.lastname: Kang
authors.affiliation: Department of Chemical Engineering, Konkuk University, Seoul 05029, Korea.
authors.forename: Jung-Kul
authors.initials: JK
authors.lastname: Lee
mesh_terms: D002151:Calorimetry; D055664:Electrochemical Techniques; D004795:Enzyme Stability / Q000187:drug effects; D004800:Enzymes, Immobilized / Q000378:metabolism*; D020600:Gluconobacter oxydans / Q000201:enzymology*; D007477:Ions; D007700:Kinetics; D007064:L-Iditol 2-Dehydrogenase / Q000302:isolation & purification / Q000378:metabolism*; D008670:Metals / Q000494:pharmacology; D062105:Molecular Docking Simulation; D009249:NADP / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D013012:Sorbitol / Q000378:metabolism; D013013:Sorbose / Q000096:biosynthesis; D013379:Substrate Specificity / Q000187:drug effects; D013816:Thermodynamics
chemical_list: D004800:Enzymes, Immobilized; D007477:Ions; D008670:Metals; D011994:Recombinant Proteins; D013012:Sorbitol; D009249:NADP; D007064:L-Iditol 2-Dehydrogenase; D013013:Sorbose