Abstract
To identify a robust NADP+ dependent formate dehydrogenase from Lactobacillus buchneri NRRL B-30929 (LbFDH) with unique biochemical properties. A new NADP+ dependent formate dehydrogenase gene (fdh) was cloned from genomic DNA of L. buchneri NRRL B-30929. The recombinant construct was expressed in Escherichia coli BL21(DE3) with 6 × histidine at the C-terminus and the purified protein obtained as a single band of approx. 44kDa on SDS-PAGE and 90kDa on native-PAGE. The LbFDH was highly active at acidic conditions (pH 4.8-6.2). Its optimum temperature was 60°C and 50°C with NADP+ and NAD+, respectively and its Tm value was 78°C. Its activity did not decrease after incubation in a solution containing 20% of DMSO and acetonitrile for 6h. The KM constants were 49.8, 0.12 and 1.68mM for formate (with NADP+), NADP+ and NAD+, respectively. An NADP+ dependent FDH from L. buchneri NRRL B-30929 was cloned, expressed and identified with its unusual characteristics. The LbFDH can be a promising candidate for NADPH regeneration through biocatalysis requiring acidic conditions and high temperatures.
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