| Abstract
| - Burkholderia cepacia AC1100 is able to usethe chlorinated compound 2,4,5-trichlorophenoxyacetic acid(2,4,5-T) as the sole source of carbon and energy. CW EPR andone-dimensional ESEEM spectroscopy studiesperformed earlier indicate the presence of a Rieske-type [2Fe-2S]cluster with two coordinated histidine residues in2,4,5-T monooxygenase from B. cepacia. Thispaper describes the application of two-dimensional ESEEM(calledHYSCORE) spectroscopy for further characterization of the nitrogenssurrounding the reduced Rieske-type cluster.The HYSCORE spectra measured at field positions in theneighborhood of the principal directions of the gtensorcontain major contributions from cross-peaks correlating the twodouble-quantum transitions from each histidinenitrogen. These allow the estimation of the diagonal components ofthe hyperfine tensors along the principal axesof the g tensor: 4.05, 3.88, and 4.01 MHz (N1) and 4.71,5.07, and 5.02 MHz (N2). Other spectral features fromthe histidine nitrogens usually have a much weaker intensity and areoccasionally observed in the spectra. HYSCOREmeasurements have been also performed with the reduced [2Fe-2S] plantferredoxin-type cluster with four cysteineligands in a ferredoxin from Porphira umbilicalis, andspectral features produced by the peptide nitrogen areobserved.Similar features also appear in the HYSCORE spectra of the Rieskecluster. Systematic differences are observedbetween 2,4,5-T monooxygenase and published results from relatedbenzene and phthalate dioxygenases that mayreflect structural and functional differences in histidine ligation andthe nitrogens of nearby amino acids in Rieske-type [2Fe-2S] clusters.
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