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Polyamine oxidases are FAD-dependent enzymes catalyzing the oxidation of polyamines at the secondary amino groups.Zea maysPAO (ZmPAO) oxidizes the carbon on the endo-side of the N5-nitrogen of spermidine (Spd) and spermine (Spm). | IFEBS Journal The structure of maize polyamine oxidase K300M mutant in complex with the natural substrates provides a snapshot of the catalytic mechanism of polyamine oxidation Annarita Fiorillo1 2 Rodolfo Federico3 Fabio Polticelli3 Alberto Boffi2 Franco Mazzei4 Massimo Di Fusco4 Andrea Ilari2 and Paraskevi Tavladoraki3 1 Department of Science and BiomedicalTechnology University of L Aquila Italy 2 CNR Institute of Molecular Biology and Pathology and Department of BiochemicalSciences Sapienza University of Rome Italy 3 Department of Biology University Roma Tre Rome Italy 4 Department of Chemistry and Drug Technologies Sapienza University of Rome Italy Keywords K300M mutant spermidine spermine X-ray structures Zea mays polyamine oxidase Correspondence A. Ilari IBPM-CNR c o Department of Biochemical Sciences University of Rome Sapienza P.le A. Moro 5 00185 Rome Italy Fax 39 064 440 062 Tel 39 064 991 0990 E-mail andrea.ilari@uniroma1.it Received 1 July 2010 revised 17 December 2010 accepted 23 December 2010 doi 10.1111 j.1742-4658.2010.08000.x Polyamine oxidases are FAD-dependent enzymes catalyzing the oxidation of polyamines at the secondary amino groups. Zea mays PAO ZmPAO oxidizes the carbon on the endo-side of the N5-nitrogen of spermidine Spd and spermine Spm . The structure of ZmPAO revealed that the active site is formed by a catalytic tunnel in which the N5 atom of FAD lies in close proximity to the K300 side chain the only active-site residue conserved in all PAOs. A water molecule HOH309 is hydrogen-bound to the amino group of K300 and mutation of this residue results in a 1400fold decrease in the rate of flavin reduction. The structural studies on the catalytically impaired ZmPAO-K300M mutant described here show that substrates are bound in an out-of-register mode and the HOH309 water molecule is absent in the enzyme-substrate complexes. Moreover K300 mutation brings about a 60 mV decrease in the FAD redox potential and a 30-fold decrease in the FAD .
