Conformational changes occurring upon reduction and NO binding in nitrite reductase from Pseudomonas aeruginosa

Area of Science:

• Biochemistry
• Structural Biology
• Enzymology

Background:

• Nitrite reductase (NiR) catalyzes nitrite reduction to nitric oxide, a crucial step in denitrification.
• Pseudomonas aeruginosa NiR (NiR-Pa) and Paracoccus denitrificans NiR (NiR-Pd) are well-studied enzymes with distinct structural features.
• Previous studies have described the oxidized and reduced forms of NiR-Pd and the oxidized form of NiR-Pa.

Purpose of the Study:

• To elucidate the structural basis of nitrite reductase activity by determining the structure of reduced NiR-Pa.
• To investigate the structural changes occurring upon reduction and nitric oxide (NO) binding in NiR-Pa.
• To compare the structural dynamics of NiR-Pa with NiR-Pd during catalysis.

Main Methods:

• X-ray crystallography was used to determine the structures of reduced NiR-Pa in both unbound and NO-bound states.
• Comparative structural analysis was performed between oxidized and reduced NiR-Pa, and between NiR-Pa and NiR-Pd.

Main Results:

• The structures of reduced NiR-Pa, both unbound and with bound NO, were determined.
• Significant conformational changes were observed in the c-heme domain of NiR-Pa upon reduction, including a loop shift and rotation of the Tyr10 side chain.
• While oxidized NiR-Pa and NiR-Pd structures differ, their reduced forms exhibit high similarity, suggesting convergence in catalytic mechanisms.

Conclusions:

• The reduction of NiR-Pa and subsequent NO binding induce specific conformational alterations, particularly around the c-heme site.
• The convergence of NiR-Pa and NiR-Pd into similar reduced conformations indicates diverse pathways leading to a conserved functional state.
• These findings provide insights into the intricate structural dynamics governing nitrite reductase catalysis.