Synergistic improvement of PQQ-dependent D-sorbitol dehydrogenase activity from Gluconobacter oxydans for the biosynthesis of miglitol precursor 6-(N-hydroxyethyl)-amino-6-deoxy-α-L-sorbofuranose.
6-(N-hydroxyethyl) amino-6-deoxy-l-sorbofuranose (6NSL) is the direct precursor of miglitol for diabetes therapy. The regio- and stereo-selective dehydrogenation offered by the membrane-bound d-sorbitol dehydrogenase (mSLDH) from Gluconobacter oxydans provides an elegant enzymatic method for 6NSL production. In this study, two subunits sldA and sldB of mSLDH were introduced into G. oxydans ZJB-605, and the specific enzyme activity of mSLDH towards NHEG was enhanced by 2.15-fold. However, the endogenous PQQ level was dramatically reduced in the recombinant strain and became a bottleneck to support the holo-enzyme activity. A combined supplementation of four amino acids (Glu, Ile, Ser, Arg) involved in biosynthesis of PQQ in conventional media effectively increased extracellular accumulation of PQQ by 1.49-fold, which further enhanced mSLDH activity by 1.33-fold. The synergic improvement of mSLDH activity provided in this study supports the superior high dehydrogenate activity towards substrate N-2-hydroxyethyl-glucamine, 184.28 g·L-1 of 6NSL was produced after a repeated bioconversion process catalyzed by the resting cells of G. oxydans/pBB-sldAB, all of which presenting a great potential of their industrial application in 6NSL biosynthesis.
['Ke|Xia|X|', 'Pan-Hong|Yu|Y|', 'Hu|Zhong-Ce|ZC|', 'Chen|Liang|L|', 'Sun|Xin-Qiang|XQ|', 'Zheng|Yu-Guo|YG|']
D017485:1-Deoxynojirimycin D000596:Amino Acids D001426:Bacterial Proteins D019149:Bioreactors D003470:Culture Media D005285:Fermentation D015870:Gene Expression D020600:Gluconobacter oxydans D007004:Hypoglycemic Agents D007064:L-Iditol 2-Dehydrogenase D045542:PQQ Cofactor D011994:Recombinant Proteins D013012:Sorbitol D013013:Sorbose
Gluconobacter oxydans Miglitol Membrane-bound D-sorbitol dehydrogenase Pyrroloquinoline quinine