Preparation of fluorinated amino acids with tyrosine phenol lyase - Effects of fluorination on reaction kinetics and mechanism of tyrosine phenol lyase and tyrosine protein kinase Csk

Abstract: Fluorinated analogues of L-tyrosine were synthesized enzymatically from the corresponding fluorinated phenols and ammonium pyruvate using tyrosine phenol-lyase. The isolated yields ranged from 0.5-2.8 g per liter (10-42% based on phenol added). The structure of 2,3,6-trifluoro-L-tyrosine hydrochloride hydrate was determined by x-ray crystallography. The fluorinated tyrosines are good substrates for the beta-elimination reaction catalysed by tyrosine phenol-lyase, except for 3,5-difluoro and the trifluorotyrosines. Fluorine at the 3-position increases the steady-state concentration of quinonoid intermediate absorbing at 500 nm compared to L-tyrosine, while fluorine at the 2-position dramatically decreases the intensity of the 500 nm absorbance peak. Reaction of tyrosine phenol-lyase with beta,beta,beta-trifluoroalanine and fluorophenols generates transient ring-fluorinated beta,beta-difluorotyrosines. 2,3,5-Trifluoro-L-tyrosine synthesized using tyrosine phenol-lyase was incorporated into a peptide substrate for tyrosine protein kinase Csk, and the resultant fluorinated peptide was found to be a good substrate for phosphorylation, suggesting that general base catalysis with early deprotonation is not involved in the reaction mechanism.