TAILIEUCHUNG - Báo cáo khoa học: Impact of alterations near the [NiFe] active site on the function of the H2 sensor from Ralstonia eutropha

In proteobacteria capable of H2oxidation under (micro)aerobic conditions, hydrogenase gene expression is often controlled in response to the availab-ility of H2. The H2 -sensing signal transduction pathway consists of a het-erodimeric regulatory [NiFe]-hydrogenase (RH), a histidine protein kinase and a response regulator. | ễFEBS Journal Impact of alterations near the NiFe active site on the function of the H2 sensor from Ralstonia eutropha Antje Gebler1 Tanja Burgdorf1 Antonio L. De Lacey2 Olaf Rudiger2 Arturo Martinez-Arias2 Oliver Lenz1 and Barbel Friedrich1 1 Institut fur Biologie Humboldt Universitat zu Berlin Germany 2 Instituto de Catalisis CSIC Madrid Spain Keywords hydrogen sensor NiFe active site regulatory hydrogenase Correspondence B. Friedrich Institut fur Biologie Humboldt Universitat zu Berlin Chausseestr. 117 10115 Berlin Germany Fax 49 30 209 38102 Tel 49 30 209 38101 E-mail Received 28 September 2006 revised 25 October 2006 accepted 31 October 2006 doi In proteobacteria capable of H2 oxidation under micro aerobic conditions hydrogenase gene expression is often controlled in response to the availability of H2. The H2-sensing signal transduction pathway consists of a heterodimeric regulatory NiFe -hydrogenase RH a histidine protein kinase and a response regulator. To gain insights into the signal transmission from the Ni-Fe active site in the RH to the histidine protein kinase conserved amino acid residues in the L0 motif near the active site of the RH large subunit of Ralstonia eutropha H16 were exchanged. Replacement of the strictly conserved Glu13 E13N E13L resulted in loss of the regulatory H2-oxidizing and D2 H exchange activities of the RH. According to EPR and FTIR analysis these RH derivatives contained fully assembled NiFe active sites and para- ortho-H2 conversion activity showed that these centres were still able to bind H2. This indicates that H2 binding at the active site is not sufficient for the regulatory function of H2 sensors. Replacement of His15 a residue unique in RHs by Asp restored the consensus of energy-linked NiFe -hydrogenases. The respective RH mutant protein showed only traces of H2-oxidizing activity whereas its D2 H -exchange activity and H2-sensing function were almost .

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