Document Detail

Molecular determinants of the cofactor specificity of ribitol dehydrogenase, a short-chain dehydrogenase/reductase.
MedLine Citation:
PMID:  22344653     Owner:  NLM     Status:  MEDLINE    
Ribitol dehydrogenase from Zymomonas mobilis (ZmRDH) catalyzes the conversion of ribitol to d-ribulose and concomitantly reduces NAD(P)(+) to NAD(P)H. A systematic approach involving an initial sequence alignment-based residue screening, followed by a homology model-based screening and site-directed mutagenesis of the screened residues, was used to study the molecular determinants of the cofactor specificity of ZmRDH. A homologous conserved amino acid, Ser156, in the substrate-binding pocket of the wild-type ZmRDH was identified as an important residue affecting the cofactor specificity of ZmRDH. Further insights into the function of the Ser156 residue were obtained by substituting it with other hydrophobic nonpolar or polar amino acids. Substituting Ser156 with the negatively charged amino acids (Asp and Glu) altered the cofactor specificity of ZmRDH toward NAD(+) (S156D, [k(cat)/K(m)(,NAD)]/[k(cat)/K(m)(,NADP)] = 10.9, where K(m)(,NAD) is the K(m) for NAD(+) and K(m)(,NADP) is the K(m) for NADP(+)). In contrast, the mutants containing positively charged amino acids (His, Lys, or Arg) at position 156 showed a higher efficiency with NADP(+) as the cofactor (S156H, [k(cat)/K(m)(,NAD)]/[k(cat)/K(m)(,NADP)] = 0.11). These data, in addition to those of molecular dynamics and isothermal titration calorimetry studies, suggest that the cofactor specificity of ZmRDH can be modulated by manipulating the amino acid residue at position 156.
Hee-Jung Moon; Manish Kumar Tiwari; Ranjitha Singh; Yun Chan Kang; Jung-Kul Lee
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't     Date:  2012-02-17
Journal Detail:
Title:  Applied and environmental microbiology     Volume:  78     ISSN:  1098-5336     ISO Abbreviation:  Appl. Environ. Microbiol.     Publication Date:  2012 May 
Date Detail:
Created Date:  2012-04-11     Completed Date:  2012-07-13     Revised Date:  2013-06-26    
Medline Journal Info:
Nlm Unique ID:  7605801     Medline TA:  Appl Environ Microbiol     Country:  United States    
Other Details:
Languages:  eng     Pagination:  3079-86     Citation Subset:  IM    
Department of Bioscience and Biotechnology, Konkuk University, Gwangjin-Gu, Seoul, Republic of Korea.
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MeSH Terms
Amino Acid Sequence
Amino Acid Substitution
Coenzymes / metabolism*
Molecular Sequence Data
Mutagenesis, Site-Directed
NAD / metabolism
NADP / metabolism*
Pentoses / metabolism
Protein Binding
Ribitol / metabolism
Sequence Homology, Amino Acid
Sugar Alcohol Dehydrogenases / genetics*,  metabolism*
Zymomonas / enzymology*
Reg. No./Substance:
0/Coenzymes; 0/Pentoses; 488-81-3/Ribitol; 53-59-8/NADP; 53-84-9/NAD; 5556-48-9/ribulose; EC 1.1.-/Sugar Alcohol Dehydrogenases; EC 2-dehydrogenase

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