title: Protein conformational changes determined by matrix-assisted laser desorption mass spectrometry.
abstract: It is shown that simultaneously to the unfolding of hen egg white lysozyme and horse heart cytochrome c the sequential conformational changes and molten globule states can be detected by the combination of proteolysis and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). This is demonstrated by the differences among the products and the time courses of native lysozyme as well as those unfolded in 1 and 3 M guanidine hydrochloride (GuHCl) when they were proteolyzed by proteinase K and analyzed by MALDI-MS. Due to the absence of disulfide bonds in the cytochrome c molecule, it is more sensitive to the disturbance of the denaturant. The partially unfolded state as detected at low concentrations of guanidine hydrochloride in our experiment resemble the molten globule state. One of the unique properties of the method described herein is to measure directly the peptide fragment liberated from proteolysis of the protein. It allows the identification of the sensitive sites susceptible to denaturation, which are subsequently cleaved by proteinase K proteolysis.
authors.affiliation: Institut für Physikalische Chemie, Universität Würzburg, Marcusstrasse 9/11, Würzburg, D-97070, Germany.
authors.forename: H H
authors.initials: HH
authors.lastname: Yang
authors.forename: X C
authors.initials: XC
authors.lastname: Li
authors.forename: M
authors.initials: M
authors.lastname: Amft
authors.forename: J
authors.initials: J
authors.lastname: Grotemeyer
mesh_terms: D000818:Animals; D001665:Binding Sites; D002645:Chickens; D003574:Cytochrome c Group / Q000737:chemistry; D019286:Endopeptidase K; D006736:Horses; D008958:Models, Molecular; D009113:Muramidase / Q000737:chemistry; D010446:Peptide Fragments / Q000302:isolation & purification; D011487:Protein Conformation; D011489:Protein Denaturation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D003574:Cytochrome c Group; D010446:Peptide Fragments; D009113:Muramidase; D019286:Endopeptidase K