Mechanisms of inhibition of phenylalanine ammonia-lyase by phenol inhibitors and phenol/glycine synergistic inhibitors

Abstract

Phenylalanine ammonia-lyase (PAL) catalyzes the β-elimination of ammonia from l-phenylalanine to trans-cinnamic acid. A study of inhibition of PAL by phenol, ortho-cresol, and meta-cresol gave mixed inhibition; para-cresol is not an inhibitor. The calculated values of K i and αK i are phenol, K i=2.1±0.5 mM and αK i=3.45±0.95 mM; ortho-cresol, K i=0.8±0.2 mM and αK i=3.4±0.2 mM; meta-cresol, K i=2.85±0.15 mM and αK i=18.5±1.5 mM. The synergistic inhibition of the same inhibitors with glycine showed a lack of inhibition with the para-cresol/glycine pair, while mixed inhibition was observed with the ortho-cresol/glycine pair ( K i=0.038±0.008 mM, αK i=0.13±0.04 mM) and phenol/glycine pair ( K i=0.014±0.003 mM, αK i=0.058±0.01 M). The meta-cresol/glycine pair gave competitive inhibition ( K i=0.36±0.076 mM). The strong synergistic inhibition observed implies that the inhibitors bind at the active site: in fact, the inhibitors used imitate the structure of the substrate. The order of synergistic inhibition is the same for the sites related to K i and αK i. These results are in agreement with the inhibitors entering two active sites located in two different subunits.