Tyrosine ammonia-lyase (TAL) is a recently described member of the aromatic amino acid lyase family, which also includes phenylalanine (PAL) and histidine ammonia-lyases (HAL). TAL is highly selective for L-tyrosine, and synthesizes 4-coumaric acid as a protein cofactor or antibiotic precursor in microorganisms. In this report, we identify a single active site residue important for substrate selection in this enzyme family. Replacing the active site residue His89 with Phe in TAL completely switched its substrate selectivity from tyrosine to phenylalanine, thereby converting it into a highly active PAL. When a corresponding mutation was made in PAL, the enzyme lost PAL activity and gained TAL activity. The discovered substrate selectivity switch is a rare example of a complete alteration of substrate specificity by a single point mutation. We also show that the identity of the amino acid at the switch position can serve as a guide to predict substrate specificities of annotated aromatic amino acid lyases in genome sequences.
Bibliographical noteFunding Information:
The authors gratefully acknowledge Dr. Gordon Louie and Dr. Joseph Noel (Salk Institute) for the communication of their TAL structural data immediately prior to publication. The authors also acknowledge Dr. Yuk Yin Sham for assistance with TAL modeling. This investigation has been supported by a grant from the David and Lucile Packard Foundation (grant 2001-18996). K.T.W. was supported by an NIGMS/NIH Biotechnology Training Grant (T32 GM08347) and the University of Minnesota Presidential Initiative on Biocatalysis.