MALDI mass spectrometry of dye-peptide and dye-protein complexes.
Immobilized sulfonate dyes are widely used for protein separation and purification, but the mode of interaction between the dye molecules and the proteins is largely unknown. Here we show that specific noncovalent dye-protein and dye-peptide complexes can be observed using MALDI mass spectrometry. We prove that the interaction is prodominantly electrostatic and that it involves protonated sites of the peptides and proteins, including the NH2 terminus, and deprotonated SO3 groups of the dyes. Furthermore, we show that MALDI-MS of such complexes with a nonacidic matrix, p-nitro-aniline, can be used to determine the number of accessible basic sites of a protein or peptide in its folded structure. Our results are in good agreement with measurements of the same property done with electrospray ionization.
Salih
B
B
Department of Chemistry, ETH Zentrum, Zürich, Switzerland.
Zenobi
R
R
D000324:Adrenocorticotropic Hormone / Q000737:chemistry; D000814:Aniline Compounds / Q000737:chemistry; D001557:Benzenesulfonates / Q000737:chemistry; D001665:Binding Sites; D001839:Bombesin / Q000737:chemistry; D004396:Coloring Agents / Q000737:chemistry*; D003574:Cytochrome c Group / Q000737:chemistry; D007328:Insulin / Q000737:chemistry; D008555:Melitten / Q000737:chemistry; D010455:Peptides / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods*; D013373:Substance P / Q000737:chemistry; D014227:Triazines / Q000737:chemistry; D014452:Ubiquitins / Q000737:chemistry
D000814:Aniline Compounds; D001557:Benzenesulfonates; D004396:Coloring Agents; D003574:Cytochrome c Group; D007328:Insulin; D010455:Peptides; D011506:Proteins; D014227:Triazines; D014452:Ubiquitins; D008555:Melitten; C008625:nitroaniline; D013373:Substance P; C012842:Cibacron Blue F 3GA; C032365:benzenesulfonic acid; D000324:Adrenocorticotropic Hormone; D001839:Bombesin