A dye-photosensitized reaction that generates stable protein-protein crosslinks.
Irradiation of fibrinogen with visible light for 30 s in the presence of 1-1000 microM fluorescein was found to crosslink fibrinogen both inter- and intramolecularly. Optimum crosslinking was achieved at dye concentrations of around 100 microM and the amount of crosslinking was shown to increase with pH. Crosslinking was inhibited in the presence of 50 microM tryptophan or tyrosine and enhanced in the presence of 5 mM histidine. Twice as much crosslinking was found to take place under anaerobic conditions. These observations are consistent with a dye-photosensitized reaction following the hydrogen abstraction pathway. The subunits of eukaryotic ribosomes and those of phosphorylase a were also crosslinked by the method described.
Department of Biochemistry and Microbiology, Scotland, United Kingdom.
A
A
Webster
D
D
Britton
A
A
Apap-Bologna
G
G
Kemp
D000596:Amino Acids / Q000032:analysis; D001777:Blood Coagulation / Q000187:drug effects; D004396:Coloring Agents; D003432:Cross-Linking Reagents; D005340:Fibrinogen / Q000032:analysis* / Q000494:pharmacology / Q000528:radiation effects; D005452:Fluoresceins; D006863:Hydrogen-Ion Concentration; D008027:Light; D010762:Phosphorylase a / Q000032:analysis; D011506:Proteins / Q000032:analysis*
D000596:Amino Acids; D004396:Coloring Agents; D003432:Cross-Linking Reagents; D005452:Fluoresceins; D011506:Proteins; D005340:Fibrinogen; D010762:Phosphorylase a