Catalysis in the nitrilase superfamily

Area of Science:

• Biochemistry
• Enzymology
• Structural Biology

Background:

• The nitrilase superfamily, named after plant and bacterial nitrilases, encompasses diverse enzyme families.
• A novel Glu-Lys-Cys catalytic triad was previously identified in the worm NitFhit active site.
• This triad was postulated as the conserved catalytic motif for the entire superfamily.

Purpose of the Study:

• To confirm the essentiality of the proposed Glu-Lys-Cys catalytic triad in the nitrilase superfamily.
• To elucidate the biochemical functions of various branches within the nitrilase superfamily.
• To expand the definition and understanding of the nitrilase superfamily.

Main Methods:

• Crystallographic analysis of the NitFhit active site.
• Experimental validation of catalytic triad residues.
• Biochemical assays to determine enzyme functions.

Main Results:

• The essentiality of the Glu-Lys-Cys catalytic triad residues was experimentally confirmed.
• Biochemical functions of additional nitrilase superfamily branches and sub-branches were specified.
• The definition of the nitrilase superfamily was expanded to include 12 families beyond nitrilases.

Conclusions:

• The Glu-Lys-Cys catalytic triad is a conserved and essential feature across the nitrilase superfamily.
• Recent experimental data validate and refine the understanding of nitrilase superfamily members and their functions.