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Mutagenesis of nitrite reductase from Pseudomonas aeruginosa : tyrosine‐10 in the c heme domain is not involved in catalysis 1
Author(s) -
Cutruzzolà Francesca,
Arese Marzia,
Grasso Sabrina,
Bellelli Andrea,
Brunori Maurizio
Publication year - 1997
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(97)00583-8
Subject(s) - nitrite reductase , chemistry , heme , enzyme , pseudomonas aeruginosa , stereochemistry , biochemistry , enzyme kinetics , nitrite , mutagenesis , tyrosine , mutant , active site , bacteria , nitrate reductase , biology , organic chemistry , nitrate , gene , genetics
In Pseudomonas aeruginosa , conversion of nitrite to NO in dissimilatory denitrification is catalyzed by the enzyme nitrite reductase (NiR), a homodimer containing a covalently bound c heme and a d 1 heme per subunit. We report the purification and characterization of the first single mutant of P. aeruginosa cd 1 NiR in which Tyr 10 has been replaced by Phe; this amino acid was chosen as a possibly important residue in the catalytic mechanism of this enzyme based on the proposal (Fülöp, V., Moir, J.W.B., Ferguson, S.J. and Hajdu, J. (1995) Cell 81, 369–377) that the topologically homologous Tyr 25 plays a crucial role in controlling the activity of the cd 1 NiR from Thiosphaera pantotropha . Our results show that in P. aeruginosa NiR substitution of Tyr 10 with Phe has no effect on the activity, optical spectroscopy and electron transfer kinetics of the enzyme, indicating that distal coordination of the Fe 3+ of the d 1 heme is provided by different side‐chains in different species.