The nitrite reductase from P. aeruginosa is a homodimer. Each protein chain (Mr 60 kDa) begins at the amino terminus with a nonstructured segment (amino acids 1-29; 1-5 are within the crystal too variable to be seen in the x-ray structure) . Next along the chain there is a domain resembling cytochrome C (amino acids 30-115), to which a C-heme is bound . An intervening segment (116-149) contains a helix (120-128, red) and connects to a beta propeller domain with a d1 heme embedded .
C-domain: The C-heme bound to this part figures the electron acceptor part of the molecule. In vitro it may be reduced by azurine oder cytochrome c551. The heme is situated in a pocket formed by hydrophobic and semipolar amino acids . The propionate groups of the heme are held in place by salt bridges . Via the sulfur atoms of two cysteines the heme is bound covalently to the protein. The iron atom is coordinated both by the heme nitrogen atoms and a histidine and a methionine . The protein topology of the C-domain closely resembles a cytochrome C. Helices are dominant features in this domain . D1-domain: In contrast to the C-domain this part of the molecule is constructed mainly from beta sheets . The quasi rotation symmetry of the order of the sheets imposes the impression of an eight bladed propeller to this domain. Very few short helices are found here . The d1 heme is bound about centered at the lower part of the beta sheet ring . This heme is not bound covalently, instead it is held by hydrophobic forces in a pocket of the protein and additionally by ionic forces via its four carboxyl groups . A pore filled with water connects one of the carboxyl groups with the surface of the protein. Polar amino acids are involved in binding the water . In the protein crystal a phosphate ion is bound to the water molecule at the exit of the channel . The fifth ligand of the iron atom in the d1 heme is the ring nitogen of his182 . The sixth ligand is not an amino acid but a hydroxyl ion . The substrate of the enzyme (NO2-) is bound by the histidine residues 327 and 369 . These amino acids are held in place by hydrogen bonds to the neighbouring leucine and lysine. The function of the hydroxyl ion is to substitute the NO after the reduction step to release it from the iron atom. The accessibility of the prostetic groups is visible in a space fillig aspect of the protein . The C-heme is clearly visible. It needs free accessibility to be able to contact its natural reaction partner cytochrome c551. The d1 heme is more strongly shielded from the environment - its low molecular weight substrate is capable to diffuse through small openings (use the mouse to find the opening!). Also to be seen is the position of the N-domain within the other monomer. The ligand Tyr10 of the d1 heme originates from the other subunit. This structure (domain swapping) contributes to the stability of the dimer. There are further intermolecular contacts between the d1-domains : hydrogen bonds connect adjacent protein strands. Chloride ions (green) form additional hydrogen bonds to amino groups. Literature: D Nurizzo et al, Structure 15 (1997) 1157-1171 Nitrite reduction Class I enzyme Table of contents . . . . 5-98 © R Bergmann If you don't see a molecular model:http://www.biologie.uni-hamburg.de/lehre/bza/1nir/e1nirm.htm
D1-domain: In contrast to the C-domain this part of the molecule is constructed mainly from beta sheets . The quasi rotation symmetry of the order of the sheets imposes the impression of an eight bladed propeller to this domain. Very few short helices are found here . The d1 heme is bound about centered at the lower part of the beta sheet ring . This heme is not bound covalently, instead it is held by hydrophobic forces in a pocket of the protein and additionally by ionic forces via its four carboxyl groups .
A pore filled with water connects one of the carboxyl groups with the surface of the protein. Polar amino acids are involved in binding the water . In the protein crystal a phosphate ion is bound to the water molecule at the exit of the channel . The fifth ligand of the iron atom in the d1 heme is the ring nitogen of his182 . The sixth ligand is not an amino acid but a hydroxyl ion . The substrate of the enzyme (NO2-) is bound by the histidine residues 327 and 369 . These amino acids are held in place by hydrogen bonds to the neighbouring leucine and lysine. The function of the hydroxyl ion is to substitute the NO after the reduction step to release it from the iron atom. The accessibility of the prostetic groups is visible in a space fillig aspect of the protein . The C-heme is clearly visible. It needs free accessibility to be able to contact its natural reaction partner cytochrome c551. The d1 heme is more strongly shielded from the environment - its low molecular weight substrate is capable to diffuse through small openings (use the mouse to find the opening!). Also to be seen is the position of the N-domain within the other monomer. The ligand Tyr10 of the d1 heme originates from the other subunit. This structure (domain swapping) contributes to the stability of the dimer. There are further intermolecular contacts between the d1-domains : hydrogen bonds connect adjacent protein strands. Chloride ions (green) form additional hydrogen bonds to amino groups. Literature: D Nurizzo et al, Structure 15 (1997) 1157-1171 Nitrite reduction Class I enzyme Table of contents . . . . 5-98 © R Bergmann If you don't see a molecular model:http://www.biologie.uni-hamburg.de/lehre/bza/1nir/e1nirm.htm
The accessibility of the prostetic groups is visible in a space fillig aspect of the protein . The C-heme is clearly visible. It needs free accessibility to be able to contact its natural reaction partner cytochrome c551. The d1 heme is more strongly shielded from the environment - its low molecular weight substrate is capable to diffuse through small openings (use the mouse to find the opening!). Also to be seen is the position of the N-domain within the other monomer. The ligand Tyr10 of the d1 heme originates from the other subunit. This structure (domain swapping) contributes to the stability of the dimer. There are further intermolecular contacts between the d1-domains : hydrogen bonds connect adjacent protein strands. Chloride ions (green) form additional hydrogen bonds to amino groups.
Literature: D Nurizzo et al, Structure 15 (1997) 1157-1171
