Unusual Fragmentation Pathways in Collagen Glycopeptides

  • Irina Perdivara
  • Lalith Perera
  • Marnisa Sricholpech
  • Masahiko Terajima
  • Nancy Pleshko
  • Mitsuo Yamauchi
  • Kenneth B. Tomer
Research Article

DOI: 10.1007/s13361-013-0624-y

Cite this article as:
Perdivara, I., Perera, L., Sricholpech, M. et al. J. Am. Soc. Mass Spectrom. (2013) 24: 1072. doi:10.1007/s13361-013-0624-y

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.

Key words

Collagen Glycopeptide Hydroxyproline Galactosylhydroxylysine Tandem mass spectrometry Fragmentation pathway Mobile proton 

Supplementary material

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Copyright information

© American Society for Mass Spectrometry (outside the USA) 2013

Authors and Affiliations

  • Irina Perdivara
    • 1
  • Lalith Perera
    • 2
  • Marnisa Sricholpech
    • 3
  • Masahiko Terajima
    • 4
  • Nancy Pleshko
    • 5
  • Mitsuo Yamauchi
    • 4
  • Kenneth B. Tomer
    • 1
  1. 1.Mass Spectrometry GroupNIH/National Institute of Environmental Health SciencesResearch Triangle ParkUSA
  2. 2.Computational Chemistry GroupNIH/National Institute of Environmental Health SciencesResearch Triangle ParkUSA
  3. 3.Faculty of Dentistry, Srinakarinwirot UniversityBangkokThailand
  4. 4.School of DentistryUniversity of North Carolina at Chapel HillChapel HillUSA
  5. 5.Department of BioengineeringTemple UniversityPhiladelphiaUSA

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