title: Protein thioacylation: 2. Reagent stability in aqueous media and thioacylation kinetics.
abstract: Several thioacylating reagents have been tested toward hydrolysis under conditions suitable for protein modifications: 20-35 degrees C and buffered solutions at pH 7.5-8.5. Aliphatic dithioesters are sufficiently stable in aqueous media at room temperature (or below) if protein modification reaction time does not exceed 24 h, whereas at 35 degrees C reaction times must be limited to a few hours. Kinetic data obtained in gelatin thioacylation at room temperature using aliphatic dithioesters and dithio acid are consistent with a second-order reaction rate with respect to amine concentration. The pH dependence of the second-order reaction rate constants indicate that dithioester reacts exclusively with the free amine form of lysine residue, whereas dithiocarboxylate ion reacts with both amine and ammonium ion, probably through a more complex mechanism. Interestingly thioacylation using dithio acids may be obtained in pH near neutrality or in slightly acidic media, thus offering protein modification possibilities at pH 5-9. Thioacylation reaction rates may be expressed as R = -(dAt/dt) = k[H3O+](-b)At2[thioacylating agent] in which At is the amine concentration at time t, constants k and b depending on the reagent nature.
authors.affiliation: Centre de Recherche, Université de Bretagne-Sud, Boulevard Flandres-Dunkerque, F-56325 Lorient, France.
authors.forename: G
authors.initials: G
authors.lastname: Levesque
authors.forename: P
authors.initials: P
authors.lastname: Arsène
authors.forename: V
authors.initials: V
authors.lastname: Fanneau-Bellenger
authors.forename: T N
authors.initials: TN
authors.lastname: Pham
mesh_terms: D000215:Acylation; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007202:Indicators and Reagents; D007700:Kinetics; D008239:Lysine / Q000737:chemistry; D011506:Proteins / Q000737:chemistry*; D013056:Spectrophotometry, Ultraviolet; D013438:Sulfhydryl Compounds / Q000737:chemistry; D013696:Temperature; D014867:Water
chemical_list: D007202:Indicators and Reagents; D011506:Proteins; D013438:Sulfhydryl Compounds; D014867:Water; D008239:Lysine