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Enzyme Nicotinamide Cofactor Specificity Reversal Guided by Automated Structural Analysis and Library Design
Jackson K B Cahn1, Sabine Brinkmann-Chen1, Frances H Arnold2
1Department of Chemistry and Chemical Engineering, California Institute of Technology, 1200 E. California Blvd., Pasadena, 91125, CA, USA.
Methods in Molecular Biology (Clifton, N.J.)
|November 25, 2017
Summary
Researchers created CSR-SALAD, a computational tool to easily design enzyme mutants. This tool helps reverse cofactor specificity for nicotinamide adenine dinucleotide (NAD) or nicotinamide adenine dinucleotide phosphate (NADP) in metabolic engineering.
Area of Science:
- Biochemistry and Molecular Biology
- Synthetic Biology
- Metabolic Engineering
Background:
- Enzyme specificity for nicotinamide adenine dinucleotide (NAD) or nicotinamide adenine dinucleotide phosphate (NADP) presents challenges in metabolic engineering.
- Switching cofactor specificity is crucial for optimizing synthetic biology applications.
Purpose of the Study:
- To develop a user-friendly computational tool, CSR-SALAD, for designing enzyme mutants.
- To simplify the process of reversing cofactor specificity in enzymes.
- To provide guidelines for optimal tool usage and laboratory application.
Main Methods:
- Development of the CSR-SALAD computational tool.
- Design of mutant libraries for reversing enzyme cofactor specificity.
- Description of laboratory protocols for applying the tool.
Main Results:
- Successful development of CSR-SALAD, an accessible tool for enzyme engineering.
- Demonstration of a simplified method for reversing NAD/NADP specificity.
- Establishment of practical laboratory application methods.
Conclusions:
- CSR-SALAD facilitates the engineering of enzymes with altered cofactor specificity.
- The tool and associated methods support advancements in metabolic engineering and synthetic biology.
- Enzyme specificity reversal is made more attainable for researchers.
