tailieunhanh - Báo cáo khoa học: The Pseudomonas aeruginosa nirE gene encodes the S-adenosyl-L-methionine-dependent uroporphyrinogen III methyltransferase required for heme d1 biosynthesis

Biosynthesis of hemed1, the essential prosthetic group of the dissimilatory nitrite reductase cytochromecd1 , requires the methylation of the tetrapyr-role precursor uroporphyrinogen III at positions C-2 and C-7. We pro-duced Pseudomonas aeruginosaNirE, a putativeS-adenosyl-l-methionine (SAM)-dependent uroporphyrinogen III methyltransferase, as a recombi-nant protein inEscherichia coliand purified it to apparent homogeneity by metal chelate and gel filtration chromatography. | ỊFEBS Journal The Pseudomonas aeruginosa nirE gene encodes the S-adenosyl-L-methionine-dependent uroporphyrinogen III methyltransferase required for heme d1 biosynthesis Sonja Storbeck1 Johannes Walther1 Judith Muller1 Vina Parmar2 Hans Martin Schiebel3 Dorit Kemken4 Thomas Dulcks4 Martin J. Warren2 and Gunhild Layer1 1 Institute of Microbiology Technische Universitat Braunschweig Germany 2 Department of Biosciences University of Kent Canterbury UK 3 Institute of Organic Chemistry Technische Universitat Braunschweig Germany 4 Institute of Organic Chemistry University of Bremen Germany Keywords heme d1 biosynthesis precorrin-2 Pseudomonas aeruginosa SAM-dependent uroporphyrinogen III methyltransferase uroporphyrinogen III Correspondence G. Layer Institute of Microbiology Technische Universitat Braunschweig Spielmannstr. 7 38106 Braunschweig Germany Fax 49 531 391 5854 Tel 49 531 391 5813 E-mail Website http ifm Received 23 June 2009 revised 10 August 2009 accepted 14 August 2009 doi Biosynthesis of heme d1 the essential prosthetic group of the dissimilatory nitrite reductase cytochrome cd1 requires the methylation of the tetrapyrrole precursor uroporphyrinogen III at positions C-2 and C-7. We produced Pseudomonas aeruginosa NirE a putative S-adenosyl-L-methionine SAM -dependent uroporphyrinogen III methyltransferase as a recombinant protein in Escherichia coli and purified it to apparent homogeneity by metal chelate and gel filtration chromatography. Analytical gel filtration of purified NirE indicated that the recombinant protein is a homodimer. NirE was shown to be a SAM-dependent uroporphyrinogen III methyltransferase that catalyzes the conversion of uroporphyrinogen III into precorrin-2 in vivo and in vitro. A specific activity of nmol of precorrin-2 h-1-mg-1 of NirE was found for the conversion of uroporphyrinogen III to precorrin-2. At high enzyme concentrations NirE catalyzed an .