A comparative kinetic study on the singlet molecular oxygen-mediated photoxidation of α- and β-chymotrypsins

: Kinetic aspects of the sensitized photooxidation of α‐ and β‐chymotrypsins have been studied at pH 6 and 8. The sensitization, employing classical O2(1Δg)‐photogenerators, such as xanthene dyes, is a kinetically intricate process because of the presence of ground state dye–protein associations and to the simultaneous participation of superoxide ion and singlet molecular oxygen [O2(1Δg)]. Both proteins, that possess the same distribution pattern of photooxidizable amino acids, suffer a pure O2(1Δg)‐mediated photodynamic attack, using the carbonylic sensitizer Perinaphthenone. Overall and reactive rate constants for the O2(1Δg)‐quenching (in the order of 108 and 107/M/s, respectively), and rates of oxygen consumption determined by time‐resolved, spectroscopic and polarographic methods indicate that α‐ and β‐chymotrypsins are less photooxidizable at pH 6, as a result of an enhancement of the O2(1Δg)‐physical quenching component. In general terms, β‐chymotrypsin exhibits the greater overall proclivity to interact with O2(1Δg), whereas structural factors, possibly evidenced by a higher exposure of the reactive tryptophan residues, impart an increased photooxidation degree to the proteins at pH 8, specially to the α‐chymotrypsin.