Structural Basis for Methine Excision by a Heme Oxygenase-like Enzyme

Area of Science:

• Biochemistry
• Enzymology
• Metalloprotein chemistry

Background:

• Heme oxygenase-like domain-containing oxidases (HDOs) are a diverse enzyme family.
• These enzymes commonly employ dinuclear metal cofactors.
• FlcD from Pseudomonas aeruginosa is involved in fluopsin C antibiotic biosynthesis.

Purpose of the Study:

• To investigate the cofactor and catalytic mechanism of FlcD.
• To determine the role of FlcD in methine excision during fluopsin C biosynthesis.
• To understand the structural basis for FlcD's unique catalytic activity.

Main Methods:

• X-ray crystallography to determine FlcD structure.
• Enzyme activity assays.
• Mössbauer spectroscopy and electron paramagnetic resonance (EPR) spectroscopy.

Main Results:

• FlcD functions as a dioxygenase catalyzing a four-electron oxidation.
• Crystal structures revealed a mononuclear iron cofactor in the active site.
• Spectroscopic analyses confirmed the mononuclear iron cofactor, coordinated by histidine and glutamate residues.
• The cofactor resembles those found in mononuclear non-heme iron enzymes.

Conclusions:

• FlcD utilizes a mononuclear iron cofactor, diverging from the typical dinuclear cofactor in HDOs.
• This finding challenges the established paradigm for HDO cofactors.
• The study provides insights into the catalytic mechanism of methine excision.
• It suggests convergent evolution among mononuclear iron-dependent enzymes.