title: Electrospray mass spectrometric study of melittin trypsinolysis by a kinetic approach.
abstract: The kinetics of tryptic digestion of melittin was studied by combined electrospray ionization time-of-flight mass spectrometry and high-performance liquid chromatography. The ratios of the kinetic constants for cleavage of the peptide bonds that are susceptible to trypsin action were determined. It is shown that trypsin does not manifest affinity for the hydrolysis of the peptide bonds inside the Arg,Lys cluster series as efficiently as it cleaves the peptide at the separately localized Lys residue. This feature demonstrates clearly the advantage of the kinetic approach to tryptic mapping of proteins. The kinetic approach allows the determination of not only discrete structural segments in protein structure but also their relative locations and their amino acid sequences. Using the melittin digests and some artificially prepared amino acids and dipeptides mixtures as models, it is shown that the presence and nature of basic amino acids predetermines the charge states of the molecules analyzed by electrospray but not the yields of their ions. The aliphatic parts of the molecules seem to be more important in determining the actual ion yields.
authors.affiliation: Institute for Cytology of the Russian Academy of Sciences, St. Petersburg, Russia.
authors.forename: E P
authors.initials: EP
authors.lastname: Mirgorodskaya
authors.forename: O A
authors.initials: OA
authors.lastname: Mirgorodskaya
authors.forename: S V
authors.initials: SV
authors.lastname: Dobretsov
authors.forename: A A
authors.initials: AA
authors.lastname: Shevchenko
authors.forename: A F
authors.initials: AF
authors.lastname: Dodonov
authors.forename: V I
authors.initials: VI
authors.lastname: Kozlovskiy
authors.forename: V V
authors.initials: VV
authors.lastname: Raznikov
mesh_terms: D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D006868:Hydrolysis; D007700:Kinetics; D013058:Mass Spectrometry; D008555:Melitten / Q000737:chemistry*; D014357:Trypsin
chemical_list: D008555:Melitten; D014357:Trypsin