Hydrogen/deuterium exchange and aggregation of a polyvaline and a polyleucine alpha-helix investigated by matrix-assisted laser desorption ionization mass spectrometry.
The membrane-associated pulmonary surfactant protein C (SP-C), containing a polyvaline alpha-helix, and a synthetic SP-C analogue with a polyleucine helix (SP-C(Leu)) were studied by hydrogen/deuterium exchange matrix-assisted laser desorption ionization (MALDI) mass spectrometry. SP-C, but not SP-C(Leu), formed abundant amyloid fibrils under experimental conditions. In CD(3)OD/D(2)O, 91:9 (v/v), containing 2 mM ammonium acetate, SP-C(Leu) and SP-C exchanged 40% of their exchangeable hydrogens within 1 min. This corresponds to exchange of labile side-chain hydrogen atoms, hydrogens on the N- and C-terminal heteroatoms, and amide hydrogen atoms in the unstructured N-terminal regions. After approximately 300 h, four exchangeable hydrogen atoms in SP-C(Leu) and 10 in SP-C remained unexchanged. During this time period the ion current corresponding to singly charged SP-C decreased to <10% of the initial value due to the formation of insoluble aggregates that are not detected by MALDI mass spectrometry. In contrast, the ion current for SP-C(Leu) was maintained over this time period, although the peptides were incubated together. In combination, hydrogen/deuterium exchange and aggregation data indicate that the polyleucine peptide refolds into a helix after opening, while the unfolded polyvaline peptide forms insoluble beta-sheet aggregates rather than refolding into a helix. The SP-C helix, but not the SP-C(Leu) helix, is thus in a metastable state, which may contribute to the recently observed tendency of SP-C and its precursor to misfold and aggregate in vivo.
Hosia
Waltteri
W
Department of Medical Biochemistry and Biophysics, Karolinska Institutet, S-171 77 Stockholm, Sweden.
Johansson
Jan
J
Griffiths
William J
WJ
D000818:Animals; D003903:Deuterium / Q000378:metabolism*; D006859:Hydrogen / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010455:Peptides / Q000378:metabolism*; D017433:Protein Structure, Secondary; D037721:Pulmonary Surfactant-Associated Protein C / Q000737:chemistry* / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013552:Swine
D010446:Peptide Fragments; D010455:Peptides; D037721:Pulmonary Surfactant-Associated Protein C; C027819:polyleucine; C036501:polyvaline; D006859:Hydrogen; D003903:Deuterium