PAVLOVÁ, Martina, Martin KLVAŇA, Andrea JESENSKÁ, Zbyněk PROKOP, Hana KONEČNÁ, Yukari SATO, M. TSUDA, Yuji NAGATA a Jiří DAMBORSKÝ. The Identification of Catalytic Pentad in the Haloalkane Dehalogenase DhmA from Mycobacterium avium N85: Reaction Mechanism and Molecular Evolution. Journal of Structural Biology. 2007, roč. 157, č. 2, s. 384-392. ISSN 1047-8477. |
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@article{721200, author = {Pavlová, Martina and Klvaňa, Martin and Jesenská, Andrea and Prokop, Zbyněk and Konečná, Hana and Sato, Yukari and Tsuda, M. and Nagata, Yuji and Damborský, Jiří}, article_number = {2}, keywords = {Catalytic Pentad; Haloalkane Dehalogenase; DhmA Mycobacterium avium N85}, language = {eng}, issn = {1047-8477}, journal = {Journal of Structural Biology}, title = {The Identification of Catalytic Pentad in the Haloalkane Dehalogenase DhmA from Mycobacterium avium N85: Reaction Mechanism and Molecular Evolution}, url = {http://loschmidt.chemi.muni.cz/peg/abstracts/jsb07.html}, volume = {157}, year = {2007} }
TY - JOUR ID - 721200 AU - Pavlová, Martina - Klvaňa, Martin - Jesenská, Andrea - Prokop, Zbyněk - Konečná, Hana - Sato, Yukari - Tsuda, M. - Nagata, Yuji - Damborský, Jiří PY - 2007 TI - The Identification of Catalytic Pentad in the Haloalkane Dehalogenase DhmA from Mycobacterium avium N85: Reaction Mechanism and Molecular Evolution JF - Journal of Structural Biology VL - 157 IS - 2 SP - 384-392 EP - 384-392 SN - 10478477 KW - Catalytic Pentad KW - Haloalkane Dehalogenase KW - DhmA Mycobacterium avium N85 UR - http://loschmidt.chemi.muni.cz/peg/abstracts/jsb07.html N2 - Haloalkane dehalogenase DhmA from Mycobacterium avium N85 showed poor expression and low stability when produced in Escherichia coli. Here we present expression DhmA in newly constructed pK4RP rhodococcal expression system in a soluble and stable form. Site-directed mutagenesis was used for the identification of a catalytic pentad, which makes up the reaction machinery of all currently known haloalkane dehalogenases. The putative catalytic triad Asp123, His279, Asp250 and the first halide-stabilizing residue Trp124 were deduced from sequence comparisons. The second stabilizing residue Trp164 was predicted from a homology model. Five point mutants in the catalytic pentad were constructed, tested for activity and were found inactive. A two-step reaction mechanism was proposed for DhmA. Evolution of different types of catalytic pentads and molecular adaptation towards the synthetic substrate 1,2-dichloroethane within the protein family is discussed. ER -
PAVLOVÁ, Martina, Martin KLVAŇA, Andrea JESENSKÁ, Zbyněk PROKOP, Hana KONEČNÁ, Yukari SATO, M. TSUDA, Yuji NAGATA a Jiří DAMBORSKÝ. The Identification of Catalytic Pentad in the Haloalkane Dehalogenase DhmA from Mycobacterium avium N85: Reaction Mechanism and Molecular Evolution. \textit{Journal of Structural Biology}. 2007, roč.~157, č.~2, s.~384-392. ISSN~1047-8477.
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