Ultraviolet photoinactivation of galactosyltransferase. Protection by substrates.
Galactosyltransferase was irreversibly inactivated upon exposure to ultraviolet light and the rate of inactivation followed apparent first-order kinetics. Significant protection against inactivation was observed in the presence of various combinations of substrates. UDPgalactose and Mn2+ together gave the most protection. Amino acid analyses revealed the loss of 1 mol of tryptophan per mol of galactosyltransferase upon ultraviolet photoinactivation. Further evidence for an essential trypotphan was provided by difference spectra and by inactivation with 2-hydroxy-5-nitrobenzyl bromide and protection against this reagent by Mn2+ and UDPgalactose. The protection by UDPgalactose and Mn2+ was greater than that provided by UDPgalactose alone. Since Mn2+ provided no protection by itself, this suggested that the formation of the galactosyltransferase-Mn2+-UDPgalactose complex caused a conformational change which was responsible for the observed protection of the essential tryptophanyl residue.
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002417:Cattle; D005260:Female; D007700:Kinetics; D008345:Manganese / Q000494:pharmacology; D008892:Milk / Q000201:enzymology; D008970:Molecular Weight; D011830:Radiation Effects; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet; D014364:Tryptophan / Q000032:analysis; D014466:Ultraviolet Rays; D014531:Uridine Diphosphate Galactose / Q000494:pharmacology*; D014539:Uridine Diphosphate Sugars / Q000494:pharmacology*
D000596:Amino Acids; D014539:Uridine Diphosphate Sugars; D014531:Uridine Diphosphate Galactose; D008345:Manganese; D014364:Tryptophan