Characterization of peptides resulting from digestion of human skin elastin with elastase.
Several pathological disorders are associated with abnormalities in elastic fibers, which are mainly composed of elastin. Understanding the biochemical basis of such disorders requires information about the primary structure of elastin. Since the acquisition of structural information for elastin is hampered by its extreme insolubility in water or any organic solvent, in this study, human skin elastin was digested with elastase to produce water-soluble peptides. Tandem mass spectrometry (MS/MS) experiments were performed using conventional electrospray ionization (ESI) and nano-ESI techniques coupled with ion trap and quadrupole time-of-flight (qTOF) mass analyzers, respectively. The peptides were identified from the fragment spectra using database searching and/or de novo sequencing. The cleavage sites of the enzyme and, for the first time, the extent and location of proline hydroxylation in human skin elastin were determined. A total of 117 peptides were identified with sequence coverage of 58.8%. It has been observed that 25% of proline residues in the sequenced region are hydroxylated. Elastase cleaves predominantly at the C-terminals of the amino acids Gly, Val, Leu, Ala, and Ile, and to a lesser extent at Phe, Pro, Glu, and Arg. Our results confirm a previous report that human skin elastin lacks amino acid sequences expressed by exon 26A.
Getie
M
M
Institute of Pharmaceutics and Biopharmaceutics, Martin Luther University Halle-Wittenberg, Halle (Saale), Germany.
Schmelzer
C E H
CE
Neubert
R H H
RH
D000595:Amino Acid Sequence; D004549:Elastin / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D008969:Molecular Sequence Data; D010196:Pancreatic Elastase / Q000378:metabolism*; D010455:Peptides / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism*; D012867:Skin / Q000737:chemistry*; D021241:Spectrometry, Mass, Electrospray Ionization; D014333:Tropoelastin / Q000737:chemistry
D010455:Peptides; D014333:Tropoelastin; D004549:Elastin; D010196:Pancreatic Elastase