Flavodiiron oxygen reductase from Entamoeba histolytica: Modulation of substrate preference by tyrosine 271 and lysine 53

Vera L. Gonçalves, João B. Vicente, Liliana Pinto, Célia V. Romão, Carlos Frazão, Paolo Sarti, Alessandro Giuffrè, Miguel Teixeira

Research output: Contribution to journalArticlepeer-review

16 Citations (Scopus)

Abstract

Flavodiiron proteins (FDPs) are a family of enzymes endowed with bona fide oxygen- and/or nitric-oxide reductase activity, although their substrate specificity determinants remain elusive. After a comprehensive comparison of available three-dimensional structures, particularly of FDPs with a clear preference toward either O2or NO, two main differences were identified near the diiron active site, which led to the construction of site-directed mutants of Tyr271and Lys53in the oxygen reducing Entamoeba histolytica EhFdp1. The biochemical and biophysical properties of these mutants were studied by UVvisible and electron paramagnetic resonance (EPR) spectroscopies coupled to potentiometry. Their reactivity with O2and NO was analyzed by stopped-flow absorption spectroscopy and amperometric methods. These mutations, whereas keeping the overall properties of the redox cofactors, resulted in increased NO reductase activity and faster inactivation of the enzyme in the reaction with O2, pointing to a role of the mutated residues in substrate selectivity.

Original languageEnglish
Pages (from-to)28260-28270
Number of pages11
JournalJournal of Biological Chemistry
Volume289
Issue number41
DOIs
Publication statusPublished - 10 Oct 2014

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