Gräwert, T. and Kaiser, J. and Zepeck, F. and Laupitz, R. and Hecht, S. and Amslinger, S. and Schramek, N. and Schleicher, E. and Weber, S. and Haslbeck, M. and Buchner, J. and Rieder, C. and Arigoni, D. and Bacher, A. and Eisenreich, W. and Rohdich, F. (2004) IspH protein of Escherichia coli: Studies on iron-sulfur cluster implementation and catalysis. J. Am. Chem. Soc. 126 (40), pp. 12847-12855.
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The ispH gene of Escherichia coli specifies an enzyme catalyzing the conversion of 1-hydroxy-2-methyl-2-(E)-butenyl diphosphate into a mixture of isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) in the nonmevalonate isoprenoid biosynthesis pathway. The implementation of a gene cassette directing the overexpression of the isc operon involved in the assembly of iron-sulfur clusters into an Escherichia coli strain engineered for ispH gene expression increased the catalytic activity of IspH protein anaerobically purified from this strain by a factor of at least 200. For maximum catalytic activity, flavodoxin and flavodoxin reductase were required in molar concentrations of 40 and 12 muM, respectively. EPR experiments as well as optical absorbance indicate the presence of a [3Fe-4S](+) cluster in IspH protein. Among 4 cysteines in total, the 36 kDa protein carries 3 absolutely conserved cysteine residues at the amino acid positions 12, 96, and 197. Replacement of any of the conserved cysteine residues reduced the catalytic activity by a factor of more than 70 000.
|Additional information (public):||Times Cited: 3|
|Institutions:||Chemistry and Pharmacy > Institut für Organische Chemie > Arbeitskreis Dr. Sabine Amslinger|
|Keywords:||Deoxyxylulose phosphate-pathway non-mevalonate pathway isoprenoid biosynthesis nonmevalonate pathway infectious-disease terpenoid biosynthesis 4fe-4s protein gene-cluster lytb protein enzyme|
|Subjects:||500 Science > 540 Chemistry & allied sciences|
|Refereed:||Yes, this version has been refereed|
|Created at the University of Regensburg:||No|
|Owner:||Dr. Sabine Amslinger|
|Deposited On:||24 Nov 2009 15:48|
|Last Modified:||07 Nov 2012 12:10|
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