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Structure-guided engineering of the coenzyme specificity of Pseudomonas fluorescens mannitol 2-dehydrogenase to enable efficient utilization of NAD(H) and NADP(H).
FEBS Lett. 2008 Jan 23; 582(2):233-7.FL

Abstract

The structure of Pseudomonas fluorescens mannitol 2-dehydrogenase with bound NAD+ leads to the suggestion that the carboxylate group of Asp(69) forms a bifurcated hydrogen bond with the 2' and 3' hydroxyl groups of the adenosine of NAD+ and contributes to the 400-fold preference of the enzyme for NAD+ as compared to NADP+. Accordingly, the enzyme with the Asp(69)-->Ala substitution was found to use NADP(H) almost as well as wild-type enzyme uses NAD(H). The Glu(68)-->Lys substitution was expected to enhance the electrostatic interaction of the enzyme with the 2'-phosphate of NADP+. The Glu(68)-->Lys:Asp(69)-->Ala doubly mutated enzyme showed about a 10-fold preference for NADP(H) over NAD(H), accompanied by a small decrease in catalytic efficiency for NAD(H)-dependent reactions as compared to wild-type enzyme.

Authors+Show Affiliations

Institute of Biotechnology and Biochemical Engineering, Graz University of Technology, Petersgasse 12, Graz, Austria.No affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

18082142

Citation

Bubner, Patricia, et al. "Structure-guided Engineering of the Coenzyme Specificity of Pseudomonas Fluorescens Mannitol 2-dehydrogenase to Enable Efficient Utilization of NAD(H) and NADP(H)." FEBS Letters, vol. 582, no. 2, 2008, pp. 233-7.
Bubner P, Klimacek M, Nidetzky B. Structure-guided engineering of the coenzyme specificity of Pseudomonas fluorescens mannitol 2-dehydrogenase to enable efficient utilization of NAD(H) and NADP(H). FEBS Lett. 2008;582(2):233-7.
Bubner, P., Klimacek, M., & Nidetzky, B. (2008). Structure-guided engineering of the coenzyme specificity of Pseudomonas fluorescens mannitol 2-dehydrogenase to enable efficient utilization of NAD(H) and NADP(H). FEBS Letters, 582(2), 233-7.
Bubner P, Klimacek M, Nidetzky B. Structure-guided Engineering of the Coenzyme Specificity of Pseudomonas Fluorescens Mannitol 2-dehydrogenase to Enable Efficient Utilization of NAD(H) and NADP(H). FEBS Lett. 2008 Jan 23;582(2):233-7. PubMed PMID: 18082142.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Structure-guided engineering of the coenzyme specificity of Pseudomonas fluorescens mannitol 2-dehydrogenase to enable efficient utilization of NAD(H) and NADP(H). AU - Bubner,Patricia, AU - Klimacek,Mario, AU - Nidetzky,Bernd, Y1 - 2007/12/17/ PY - 2007/09/11/received PY - 2007/12/03/revised PY - 2007/12/04/accepted PY - 2007/12/18/pubmed PY - 2008/4/2/medline PY - 2007/12/18/entrez SP - 233 EP - 7 JF - FEBS letters JO - FEBS Lett. VL - 582 IS - 2 N2 - The structure of Pseudomonas fluorescens mannitol 2-dehydrogenase with bound NAD+ leads to the suggestion that the carboxylate group of Asp(69) forms a bifurcated hydrogen bond with the 2' and 3' hydroxyl groups of the adenosine of NAD+ and contributes to the 400-fold preference of the enzyme for NAD+ as compared to NADP+. Accordingly, the enzyme with the Asp(69)-->Ala substitution was found to use NADP(H) almost as well as wild-type enzyme uses NAD(H). The Glu(68)-->Lys substitution was expected to enhance the electrostatic interaction of the enzyme with the 2'-phosphate of NADP+. The Glu(68)-->Lys:Asp(69)-->Ala doubly mutated enzyme showed about a 10-fold preference for NADP(H) over NAD(H), accompanied by a small decrease in catalytic efficiency for NAD(H)-dependent reactions as compared to wild-type enzyme. SN - 0014-5793 UR - https://www.unboundmedicine.com/medline/citation/18082142/Structure_guided_engineering_of_the_coenzyme_specificity_of_Pseudomonas_fluorescens_mannitol_2_dehydrogenase_to_enable_efficient_utilization_of_NAD_H__and_NADP_H__ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0014-5793(07)01255-0 DB - PRIME DP - Unbound Medicine ER -