Light-induced binding of riboflavin to lysozyme

Abstract

The photodynamic inactivation of lysozyme in air saturated H2O and D2O (phosphate buffer 0.05 M, pH 7.0) in the presence of methylene blue and riboflavin has been studied. When H2O was replaced by D2O a great increase in the rate of photoinactivation of lysozyme was observed. This finding, together with the fact that photooxidation is inhibited by singlet oxygen quenchers like NaN3, suggests that these reactions occur via a singlet oxygen mechanism. During the course of the studies of the riboflavin sensitized photoinactivation of lysozyme, it was found that riboflavin is strongly bound to the enzyme as a result of illumination. This finding would explain the higher quantum yield observed when riboflavin is used, although this dye is bleached during irradiation.

Triclosan–lysozyme complex: A promising antimicrobial macromolecule stable against photooxidative damage

“…1A lower and 1B upper) likely to be the direct reaction between 3 RF * and tryptophan residues in the enzyme. The interaction between LZ and RF upon illumination generates LZ-RF adduct and photooxidation of LZ might be sensitized by RF bound to the protein and in solution (Silva & Gaule, 1977). Ferrer & Silva (1984) noted that reduction and carboxymethylation of the four disulfide bonds as well as the chemical modification of the Tyr residues and the photochemical alteration of the His residue in lysozyme, do not affect the formation of the photo-induced LZ-RF bond.…”

Rate constants studies of the dye-sensitized photoinactivation of lysozyme

“…This result suggests that the photooxidation mechanism depends also on the wavelength. Recently, presence of activated oxygen in the photodynamic effect of this enzyme with methylene blue or riboflavin has also been detected (Silva and Gaule, 1977). Recently, presence of activated oxygen in the photodynamic effect of this enzyme with methylene blue or riboflavin has also been detected (Silva and Gaule, 1977).…”

“…Higher quantum yields for photoinactivation of lysozyme sensitized by riboflavin in relation to those obtained in the presence of methylene blue (Shugar, 1951;Spikes, 1969, 1970;Silva et al, 1974) have been interpreted through the photoinduced complex obtained on irradiating lysozyme in the presence of riboflavin (Silva and Gaule, 1977).…”

“…Notwithstanding the large number of papers published for many years about the sensitized photooxidation of lysozyrne (Shugar, 1959;Galiazzo et al, 1968;Hopkins and Spikes, 1970;Schmidt and Rosenkranz, 1972;Kepka and Grossweiner, 1973;Churakova et al, 1973;Risi et al, 1973;Silva et al, 1974;Silva and Gaule, 1977), little is known about the reaction mechanism of the photodynamic effect on this enzyme. Spikes and Glad (1964) have postulated that sensitized photoinactivation of trypsin in the presence of methylene blue or riboflavin-5'-phosphate (FMN), could be carried out through different reaction mechanism.…”

Isolation and photo-oxidation of lysozyme fragments

“…Photo-oxidation of lysozyme dye-sensitized by riboflavin has been studied in detail (Risi et al 1973;Silva et al 1974Silva et al , 1977Silva 1979), and can be summarized as follows: His and Trp residues present in the enzyme are selectively photo-oxidized. The only His residue is destroyed according to a very well defined first-order kinetic, whereas the destruction of the six Trp residues shows an hyperbolical pathway in a semi-logarithmic plot.…”