This chapter reviews the design and analysis of mutated proteins, particularly regarding their importance in understanding fundamentals of enzymic catalysis. Site-specific mutagenesis has proved to be a powerful tool; it has become the principal means of dissecting the active site of an enzyme to identify and quantitate the role of specific amino acids in ligand binding, specificity, and catalysis. Some of the more noteworthy triumphs are the unambiguous identification of active-site residues, the evaluation of hydrophobic and hydrogen bond strengths, the quantitation of electrostatic contributions toward catalysis, the definition of active sites at subunit interfaces, the evaluation and improvement of enzyme stability, and dissection of the subtle interplay between substrate binding and transition state stabilization. In more and more cases, mutagenesis has extended the crystallographic view of the active site by revealing the importance of movement of remote residues into the catalytic process, concomitantly verifying the transition state theory that underpins rationale for enzymic catalysis.
All Science Journal Classification (ASJC) codes
- Molecular Biology