Pseudomonas aeruginosa cytochrome C(551): probing the role of the hydrophobic patch in electron transfer.
Cytochrome c(551) from Pseudomonas aeruginosa is a monomeric redox protein of 82 amino-acid residues, involved in dissimilative denitrification as the physiological electron donor of cd(1) nitrite reductase. The distribution of charged residues on the surface of c(551) is very anisotropic: one side is richer in acidic residues whereas the other shows a ring of positive side chains, mainly lysines, located at the border of an hydrophobic patch which surrounds the heme crevice. In order to map in cytochrome c(551) the surface involved in electron transfer, we have introduced specific mutations in three residues belonging to the hydrophobic patch, namely Val23-->Asp, Pro58-->Ala and Ile59-->Glu. The effect of these mutations was analyzed studying both the self-exchange rate and the electron-transfer activity towards P. aeruginosa cd(1) nitrite reductase, the physiological partner and P. aeruginosa azurin, a copper protein often used as a model redox partner in vitro. Our results show that introduction of a negative charge in the hydrophobic patch severely hampers both homonuclear and heteronuclear electron transfer.
Cutruzzolà
Francesca
F
Dipartimento di Scienze Biochimiche, Università di Roma La Sapienza, P. le A. Moro 5, 00185 Rome, Italy. francesca.cutruzzola@uniroma1.it
Arese
Marzia
M
Ranghino
Graziella
G
van Pouderoyen
Gertie
G
Canters
Gerard
G
Brunori
Maurizio
M
D001400:Azurin / Q000737:chemistry; D001426:Bacterial Proteins; D003574:Cytochrome c Group / Q000737:chemistry* / Q000378:metabolism; D007700:Kinetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D009572:Nitrite Reductases / Q000737:chemistry / Q000378:metabolism; D019906:Nuclear Magnetic Resonance, Biomolecular; D010084:Oxidation-Reduction; D011487:Protein Conformation; D011550:Pseudomonas aeruginosa / Q000201:enzymology*; D013499:Surface Properties
D001426:Bacterial Proteins; D003574:Cytochrome c Group; D001400:Azurin; C017167:cytochrome C(551); D009572:Nitrite Reductases