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- Regulatory role of cysteines in (2R, 3R)-butanediol dehydrogenase BdhA of Bacillus velezensis strain GH1-13
- Yunhee Choi , Yong-Hak Kim
- J. Microbiol. 2022;60(4):411-418. Published online March 14, 2022
- DOI: https://doi.org/10.1007/s12275-022-2018-y
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Abstract
- Bacillus velezensis strain GH1-13 contains a (2R,3R)-butanediol dehydrogenase (R-BDH) BdhA which converts acetoin to R-BD reversibly, however, little is known about its regulatory cysteine and biological significance. We performed sitedirected mutation of three cysteines in BdhA. The C37S mutant had no enzyme activity and the C34S and C177S mutants differed from each other and wild type (WT). After zinc affinity chromatography, 1 mM ZnCl2 treatment resulted in a 3-fold enhancement of the WT activity, but reduced activity of the C34S mutant by more than 2 folds compared to the untreated ones. However, ZnCl2 treatment did not affect the activity of the C177S mutant. Most of the double and triple mutant proteins (C34S/C37S, C34S/C177S, C37S/C177S, and C34S/C37S/C177S) were aggregated in zinc resins, likely due to the decreased protein stability. All of the purified WT and single mutant proteins increased multiple intermolecular disulfide bonds in the presence of H2O2 as the buffer pH decreased from 7.5 to 5.5, whereas an intramolecular disulfide bond of cysteine 177 and another cysteine in the CGIC motif region was likely formed at pH higher than pKa of 7.5. When pH varied, WT and its C34S or C177S mutants reduced acetoin to R-BD at the optimum pH 5.5 and oxidized R-BD to acetoin at the optimum pH 10. This study demonstrated that cysteine residues in BdhA play a regulatory role for the production of acetoin and R-BD depending on pH as well as metal binding and oxidative stress.
- Changpingibacter yushuensis gen. nov., sp. nov., isolated from fluvial sediment in Qinghai Tibet Plateau of China
- Yifan Jiao , Sihui Zhang , Jing Yang , Xin-He Lai , Kui Dong , Yanpeng Cheng , Mingchao Xu , Wentao Zhu , Shan Lu , Dong Jin , Ji Pu , Ying Huang , Liyun Liu , Suping Wang , Jianguo Xu
- J. Microbiol. 2022;60(2):147-155. Published online January 7, 2022
- DOI: https://doi.org/10.1007/s12275-022-1199-8
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Abstract
- Two facultatively anaerobic, short rod-shaped, non-motile,
Gram-stain-positive, unknown bacterial strains (JY-X040T
and JY-X174) were isolated from fluvial sediments of Tongtian
River in Yushu Tibetan Autonomous Prefecture, Qinghai
province, China. Cells formed translucent, gray, round and
convex colonies, with a diameter of less than 0.5 mm after 5
days of incubation at 30°C on brain heart infusion-5% sheep
blood agar. The 16S rRNA gene sequence similarity between
strain JY-X040T and Fudania jinshanensis 313T is 93.87%.
In the four phylogenetic trees constructed based on the 16S
rRNA gene and 423 core genes, the two isolates form an independent
branch, phylogenetically closest to F. jinshanensis
313T, but could not be classified as a member of the genus
Fudania or any other genus of the family Arcanobacteriaceae.
The DNA G + C content of strain JY-X040T was 57.8%. Calculation
results
of average nucleotide identity, digital DNADNA hybridization value and amino acid identity between strain JY-X040T and F. jinshanensis 313T are 69.9%, 22.9%, and 64.1%. The major cellular fatty acids were C16:0 (23%) and C18:1ω9c (22%). The cell-wall peptidoglycan type was A5α (L-Lys-L-Ala-L-Lys-D-Glu). The polar lipids comprised diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, phosphatidylinositol mannoside and four unidentified components. The whole-cell sugars contained rhamnose and ribose. MK-10(H4) was the sole respiratory quinone. The minimum inhibitory concentration of streptomycin was 32 μg/ml. All physiological, biochemical, chemotaxonomic and genomic characteristics support that strains JY-X040T and JY-X174 represent members of a novel species in a new genus, Changpingibacter yushuensis gen. nov., sp. nov. The type strain is JY-X040T (GDMCC 1.1996T = KCTC 49514T).
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