title: Unusual fragmentation pathways in collagen glycopeptides.
abstract: Collagens are the most abundant glycoproteins in the body. One characteristic of this protein family is that the amino acid sequence consists of repeats of three amino acids -(X-Y-Gly)n. Within this motif, the Y residue is often 4-hydroxyproline (HyP) or 5-hydroxylysine (HyK). Glycosylation in collagen occurs at the 5-OH group in HyK in the form of two glycosides, galactosylhydroxylysine (Gal-HyK) and glucosyl galactosylhydroxylysine (GlcGal-HyK). In collision induced dissociation (CID), collagen tryptic glycopeptides exhibit unexpected gas-phase dissociation behavior compared to typical N- and O-linked glycopeptides (i.e., in addition to glycosidic bond cleavages, extensive cleavages of the amide bonds are observed). The Gal- or GlcGal- glycan modifications are largely retained on the fragment ions. These features enable unambiguous determination of the amino acid sequence of collagen glycopeptides and the location of the glycosylation site. This dissociation pattern was consistent for all analyzed collagen glycopeptides, regardless of their length or amino acid composition, collagen type or tissue. The two fragmentation pathways-amide bond and glycosidic bond cleavage-are highly competitive in collagen tryptic glycopeptides. The number of ionizing protons relative to the number of basic sites (i.e., Arg, Lys, HyK, and N-terminus) is a major driving force of the fragmentation. We present here our experimental results and employ quantum mechanics calculations to understand the factors enhancing the labile character of the amide bonds and the stability of hydroxylysine glycosides in gas phase dissociation of collagen glycopeptides.
authors: Mass Spectrometry Group, NIH/National Institute of Environmental Health Sciences, Research Triangle Park, NC, USA. perdivarai@niehs.nih.gov
authors: Irina
authors: I
authors: Perdivara
authors: Lalith
authors: L
authors: Perera
authors: Marnisa
authors: M
authors: Sricholpech
authors: Masahiko
authors: M
authors: Terajima
authors: Nancy
authors: N
authors: Pleshko
authors: Mitsuo
authors: M
authors: Yamauchi
authors: Kenneth B
authors: KB
authors: Tomer
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D003094:Collagen / Q000737:chemistry* / Q000235:genetics; D006020:Glycopeptides / Q000737:chemistry* / Q000235:genetics; D006031:Glycosylation; D006901:Hydroxylysine / Q000031:analogs & derivatives / Q000737:chemistry; D008969:Molecular Sequence Data; D015394:Molecular Structure; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics; D011789:Quantum Theory; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014357:Trypsin
chemical_list: D006020:Glycopeptides; D010446:Peptide Fragments; D006901:Hydroxylysine; C001295:glucosylgalactosylhydroxylysine; C007817:galactosylhydroxylysine; D003094:Collagen; D014357:Trypsin
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