| Description |
Haloalkane dehalogenases are microbial enzymes capable to cleave carbon-halogen bond in halogenated aliphatic compounds. There is a growing interest in these enzymes because of their potential use in bioremediation, as industrial biocatalysts, or as biosensors. Structurally, haloalkane dehalogenases belong to the a/b-hydrolase fold superfamily. Best studied haloalkane dehalogeneses are DhlA from Xanthobacter autotrophicus GJ10 [1], DhaA from Rhodococcus rhodochrous NCIMB 13064 [2] and LinB from Sphingomonas paucimobilis UT26 [3]. Crystal structures of these haloalkane dehalogenases are known. There is a number of DNA sequences which are expected to code haloalkane dehalogenases. This expectation is based on sequential similiraties with known haloalkane dehalogenases: DhlA, DhaA and LinB. Protein Rv2579 from Mycobacterium tuberculosis H37Rv has 68 % sequence similarity with LinB. Gene rv2579 was cloned to Escherichia coli, protein Rv2579 was overexpressed and purified to homogeneity. Haloalkane dehalogenase activity was confirmed with pure Rv2579 enzyme. Increasing number of haloalkane dehalogenase group members demands characterisation of each enzyme into more detail and substrate specificity is an important property to be evaluated Steady-state catalytic constants are being determined for the set of 31 different substrates and all currently known haloalkane dehalogenases. Multivariate statistical method - Principal component analysis [4] - will be applied on data obtained from the substrate specificity testing and the result will show structure- funcion relationships within the group of haloalkane dehalogenases. Amalgamation of structural and funcional knowledge based on in-depth studies of family of enzymes, like haloalkane dehalogenases, could bring an important and interesting knowledge into protein science. References: 1. Verschueren, K.H.G., Seljee, F., Rozeboom, H.J., Kalk, K.H., Dijkstra, B.W.: Crystallographic analysis of the catalytic mechanism of haloalkane dehalogenase. Nature, 1993. 363: p. 693-698. 2. Newman, J., Peat, T.S., Richard, R., Kan, L., Swanson, P.E., Affholter, J.A., Holmes, I.H., Schindler, J.F., Unkefer, C.J., Terwilliger, T.C.: Haloalkane dehalogenase: structure of a Rhodococcus enzyme. Biochemistry, 1999. 38: p. 16105-16114. 3. Marek, J., Vevodova, J., Kuta-Smatanova, I., Nagata, Y., Svensson, L.A., Newman, J., Takagi, M., Damborsky, J.: Crystal structure of the haloalkane dehalogenase from Sphingomonas paucimobilis UT26. Biochemistry, 2000. 39: p. 14082-14086. 4. Wold, S., Esbensen, K., Geladi, P.: Principal Component Analysis. Chemometrics and Intelligent Laboratory Systems, 1987. 2: p. 37-52.
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