Location: Renewable Product Technology ResearchTitle: Discovery of an acidic, thermostable and highly NADP+ dependent formate dehydrogenase from Lactobacillus buchneri NRRL B-30929
|ALPDAGTAS, SAADET - Yuzuncu Yil Centennial University|
|YÜCEL, SEVIL - Yildiz Technical University|
|KAPKAÇ, HANDAN - Anadolu Universtiy|
|BINAY, BARIS - Gebze Technical University|
Submitted to: Biotechnology Letters
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/15/2018
Publication Date: 5/18/2018
Citation: Alpdagtas, S., Yücel, S., Kapkaç, H.A., Liu, S., Binay, B. 2018. Discovery of an acidic, thermostable and highly NADP+ dependent formate dehydrogenase from Lactobacillus buchneri NRRL B-30929. Biotechnology Letters. 40(7): 1135-1147. doi: 10.1007/s10529-018-2568-6.
Interpretive Summary: Formate dehydrogenases are a group of enzymes that are frequently used in industry to improve the efficiency of chemical and pharmaceutical drug syntheses using biotechnological approaches. The use of these enzymes is often limited by a rapid breakdown from extreme environmental conditions, which results in a loss of activity; and their inability to efficiently use a compound called NADPH that is required for many types of synthetic reactions. In this work, we identified a novel formate dehydrogenase enzyme from a bacterium Lactobacillus buchneri that we previously obtained from a fuel ethanol production facility. This enzyme displayed tolerance to high temperatures, industrial organic solvents, and acidic conditions. Furthermore, this enzyme was able to efficiently use NADPH in the conversion reactions. The results of this study suggest that this enzyme may be ideal for industrial biosynthetic applications and could significantly reduce the production costs through improved efficiency.
Technical Abstract: Objectives: To identify a robust NADP+ dependent formate dehydrogenase from Lactobacillus buchneri NRRL B-30929 (LbFDH) with unique biochemical properties. Results: 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 x histidine at the C-terminus and the purified protein obtained as a single band of approx. 44 kDa on SDS-PAGE and 90 kDa on native-PAGE. The LbFDH was highly active at acidic conditions (pH 4.8–6.2). Its optimum temperature was 60ºC and 50ºC with NADP+ and NAD+, respectively and its Tm value was 78ºC. Its activity did not decrease after incubation in a solution containing 20% of DMSO and acetonitrile for 6 h. The KM constants were 49.8, 0.12 and 1.68 mMfor formate (with NADP+), ADP+ and NAD+, respectively. Conclusions: 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.