1 National Centre for Biomolecular Research,
Masaryk University, Kotlarska 2, 611 37 Brno, Czech Republic
2 Department of Environmental Life Sciences,
Graduate School of Life Sciences, Tohoku University, 2-1-1 Katahira, Sendai
980-8577, Japan
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..
Protein DbjA from Bradyrhizobium japonicum USDA110 has 41 % sequence
similarity with LinB. Haloalkane dehalogenase activity of DbjA was confirmed
with heterogously overexperessed pure 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.
* To whom correspondence may be addressed: Fax: 420-5-41129506; E-mail: zbynek@chemi.muni.cz.