Characterization of whole fibril-forming collagen proteins of types I, III, and V from fetal calf skin by infrared matrix-assisted laser desorption ionization mass spectrometry

Fibril-forming collagen proteins of the types 1, 111, and V were extracted from fetal calf skin, purified by differential salt precipitation, and analyzed by infrared matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (IR-MALDI-TOF-MS). Glycerol was used as liquid IR-MALDI matrix. Noncovalently bound triple helices of the types I and V were detected from the NaCl precipitate. After heating at 43degreesC for 10 min, resulting in the dissociation of the triple helix, the single a.-chain subunits were detected. For type 1, mass spectra acquired from molecular sieve chromatography fractions revealed the presence of further substructures of dimeric type and of supramolecular complexes up to the tetramer. Triple helices of type 111, stabilized by covalent disulfide bonds, were detected from the total protein precipitate also after heat treatment. For type 111, even hexamer and nonamer structures with molecular weights close to 600 and 900 kDa were recorded. For comparison, ultraviolet (UV-)MALDI-MS measurements with 2,5-dihydroxybenzoic acid as matrix were carried out with some of the samples. Here, only the single a-chains were detected with significantly reduced sensitivity.