[KSCI] Korea Science Citation Index Service

http://dx.doi.org/10.5483/BMBRep.2012.45.8.064

Functional roles of Tryptophan residues in diketoreductase from Acinetobacter baylyi

Huang, Yan (State Key Laboratory of Natural Medicines and Laboratory of Chemical Biology, China Pharmaceutical University)
Lu, Zhuo (State Key Laboratory of Natural Medicines and Laboratory of Chemical Biology, China Pharmaceutical University)
Ma, Min (State Key Laboratory of Natural Medicines and Laboratory of Chemical Biology, China Pharmaceutical University)
Liu, Nan (State Key Laboratory of Natural Medicines and Laboratory of Chemical Biology, China Pharmaceutical University)
Chen, Yijun (State Key Laboratory of Natural Medicines and Laboratory of Chemical Biology, China Pharmaceutical University)

Publication Information

BMB Reports / v.45, no.8, 2012 , pp. 452-457 More about this Journal

Abstract

Diketoreductase (DKR) from Acinetobacter baylyi contains two tryptophan residues at positions 149 and 222. Trp-149 and Trp-222 are located along the entry path of substrate into active site and at the dimer interface of DKR, respectively. Single and double substitutions of these positions were generated to probe the roles of tryptophan residues. After replacing Trp with Ala and Phe, biochemical and biophysical characteristics of the mutants were thoroughly investigated. Enzyme activity and substrate binding affinity of W149A and W149F were remarkably decreased, suggesting that Trp-149 regulates the position of substrate at the binding site. Meanwhile, enzyme activity of W222F was increased by 1.7-fold while W222A was completely inactive. In addition to lower thermostability of Trp-222 mutants, molecular modeling of the mutants revealed that Trp-222 is vital to protein folding and dimerization of the enzyme.

Keywords

Diketoreductase; Enzyme kinetics; Hydrophobic interaction; Site-directed mutagenesis; Tryptophan;

Citations & Related Records

Times Cited By Web Of Science : 0 (Related Records In Web of Science)

Reference

1	Balchin, D., Fanucchi, S., Achilonu, I., Adamson, R. J., Burke, J., Fernandes, M., Gildenhuys, S. and Dirr, H. W. (2010) Stability of the domain interface contributes towards the catalytic function at the H-site of class alpha glutathione transferase A1-1. Biochim. Biophys. Acta. 1804, 2228-2233. DOI ScienceOn
2	Wu, X. R., Wang, Y. C., Ju, J. M., Chen, C., Liu, N. and Chen, Y. J. (2009) Enantioselective synthesis of ethyl (S)-2-hydroxy-4-phenylbutyrate by recombinant diketoreductase. Tetrahedron: Asymmetry 20, 2504-2509. DOI ScienceOn
3	Lee, J., Lee, K. and Shin, S. (2000) Theoretical studies of the response of a protein structure to cavity-creating mutations. Biophys. J. 78, 1665-1671. DOI ScienceOn
4	Hong, H., Park, S., Jimenez, R. H., Rinehart, D. and Tamm, L. K. (2007) Role of aromatic side chains in the folding and thermodynamic stability of integral membrane proteins. J. Am. Chem. Soc. 129, 8320-8327. DOI ScienceOn
5	Espinoza-Fonseca, L. M., Wong-Ramirez, C. and Trujillo- Ferrara, J. G. (2010) Tyr74 is essential for the formation, stability and function of Plasmodium falciparum triosephosphate isomerase dimer. Arch. Biochem. Biophys. 494, 46-57. DOI ScienceOn
6	Eriksson, A. E., Baase, W. A., Zhang, X. J., Heinz, D. W., Blaber, M., Baldwin, E. P. and Matthews, B. W. (1992) Response of a protein structure to cavity-creating mutations and its relation to the hydrophobic effect. Science 255, 178-183. DOI
7	Myers, D. P., Jackson, L. K., Ipe, V. G., Murphy, G. E. and Phillips, M. A. (2001) Long-range interactions in the dimer interface of ornithine decarboxylase are important for enzyme function. Biochemistry 40, 13230-13236. DOI ScienceOn
8	Groemping, Y. and Hellmann, N. (2005) Spectroscopic methods for the determination of protein interactions; in Current protocols in protein science, Coligan, J. E., Dunn, B. M., Ploegh, H. L., Speicher, D. W. and Wingfield, P. T. (eds.), pp. 20.8.1-20.8.27, John Wiley & Sons, Inc, New York, USA.
9	Lustig, D. B., Kempt, C., Alam, S., Clancy, J., Yee, J. and Rafferty, S. P. (2011) Mutation of conserved tryptophan residues at the dimer interface of Staphylococcus aureus nitric oxide synthase. Arch. Biochem. Biophys. 506, 165-172. DOI ScienceOn
10	Lee, L. C., Chou, Y. L., Chen, H. H., Lee, Y. L. and Shaw, J. F. (2009) Functional role of a non-active site residue Trp(23) on the enzyme activity of Escherichia coli thioesterase I/protease I/lysophospholipase L(1). Biochim. Biophys. Acta. 1794, 1467-1473. DOI ScienceOn
11	Hu, H. Y., Wu, M. C., Fang, H. J., Forrest, M. D., Hu, C. K., Tsong, T. Y. and Chen, H. M. (2010) The role of tryptophan in staphylococcal nuclease stability. Biophys. Chem. 151, 170-177. DOI ScienceOn
12	Lanzarotti, E., Biekofsky, R. R., Estrin, D. A., Marti, M. A. and Turjanski, A. G. (2011) Aromatic-aromatic interactions in proteins: beyond the dimer. J. Chem. Inf. Model. 51, 1623-1633. DOI ScienceOn
13	Allocati, N., Masulli, M., Pietracupa, M., Favaloro, B., Federici, L. and Di Ilio, C. (2005) Contribution of the two conserved tryptophan residues to the catalytic and structural properties of Proteus mirabilis glutathione S-transferase B1-1. Biochem. J. 385, 37-43. DOI ScienceOn
14	Bourbon-Freie, A., Dub, R. E., Xiao, X. and Lowe, M. E. (2009) Trp-107 and trp-253 account for the increased steady state fluorescence that accompanies the conformational change in human pancreatic triglyceride lipase induced by tetrahydrolipstatin and bile salt. J. Biol. Chem. 284, 14157-14164. DOI ScienceOn
15	Pace, C. N. (1995) Evaluating contribution of hydrogen bonding and hydrophobic bonding to protein folding. Methods Enzymol. 259, 538-554. DOI
16	Burley, S. K. and Petsko, G. A. (1985) Aromatic-aromatic interaction: a mechanism of protein structure stabilization. Science 229, 23-28. DOI
17	Wu, X. R., Jiang, J. P. and Chen, Y. J. (2011) Correlation between intracellular cofactor concentrations and biocatalytic efficiency: co-expression of diketoreductase and glucose dehydrogenase for the preparation of chiral diol for statin drugs. ACS Catal. 1, 1661-1664. DOI ScienceOn
18	Wu, X., Wang, L., Wang, S. and Chen, Y. (2010) Stereoselective introduction of two chiral centers by a single diketoreductase: an efficient biocatalytic route for the synthesis of statin side chains. Amino. Acids. 39, 305-308. DOI
19	Wu, X., Chen, C., Liu, N. and Chen, Y. (2011) Preparation of ethyl 3R,5S-6-(benzyloxy)-3,5-dihydroxy-hexanoate by recombinant diketoreductase in a biphasic system. Bioresour. Technol. 102, 3649-3652. DOI ScienceOn
20	Wu, X., Chen, C., Liu, N. and Chen, Y. (2010) Two step process for diketo-reduction catalyzed by diketoreductase. J. Chin. Pharma. Uni. 41, 408-413.
21	Huang, Y., Liu, N., Wu, X. R. and Chen, Y. J. (2010) Dehydrogenases/reductases for the synthesis of chiral pharmaceutical intermediates. Curr. Org. Chem. 14, 1447-1460. DOI ScienceOn
22	Lu, M. L., Cao, X., Yang, X., Zheng, H., Liu, N., Jiang, Y., Lin, D. H. and Chen, Y. J. (2010) A diketoreductase exhibits unique renaturation profile from thermal-induced protein unfolding. Amino. Acids. 39, 609-613. DOI
23	Huang, Y., Lu, Z., Liu, N. and Chen, Y. (2012) Identification of important residues in diketoreductase from Acinetobacter baylyi by molecular modeling and site-directed mutagenesis. Biochimie. 94, 471-478. DOI ScienceOn
24	Shih, C., Museth, A. K., Abrahamsson, M., Blanco- Rodriguez, A. M., Di Bilio, A. J., Sudhamsu, J., Crane, B. R., Ronayne, K. L., Towrie, M., Vlcek, A. Jr., Richards, J. H., Winkler, J. R. and Gray, H. B. (2008) Tryptophan-accelerated electron flow through proteins. Science 320, 1760-1762. DOI ScienceOn
25	Lehrer, S. S. (1971) Solute perturbation of protein fluorescence. The quenching of the tryptophyl fluorescence of model compounds and of lysozyme by iodide ion. Biochemistry 10, 3254-3263. DOI ScienceOn
26	Wu, X. R., Liu, N., He, Y. M. and Chen, Y. J. (2009) Cloning, expression, and characterization of a novel diketoreductase from Acinetobacter baylyi. Acta. Biochim. Biophys. Sin. (Shanghai) 41, 163-170. DOI ScienceOn
27	Wu, X. R., Liu, N., He, Y. M. and Chen, Y. J. (2008) A bacterial enzyme catalyzing double reduction of a ${\beta},\;{\delta}$ -diketo ester with unprecedented stereoselectivity. Nature Precedings, http://hdl.handle.net/10101/npre.12008.11697.10101.
28	Blatt, E., Chatelier, R. C. and Sawyer, W. H. (1986) Effects of quenching mechanism and type of quencher association on Stern-Volmer plots in compartmentalized systems. Biophys. J. 50, 349-356. DOI ScienceOn