4.8 Article

Tandem mass spectrometry for structural characterization of proline-rich proteins: Application to salivary PRP-3

Journal

ANALYTICAL CHEMISTRY
Volume 74, Issue 16, Pages 4124-4132

Publisher

AMER CHEMICAL SOC
DOI: 10.1021/ac0255835

Keywords

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Funding

  1. NCRR NIH HHS [P41 RR 10888] Funding Source: Medline
  2. NIDCR NIH HHS [R01 DE 07652, R01 DE 05672] Funding Source: Medline

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Proline-rich proteins (PRPs), including collagens, complement 1q, and salivary PRPs, are unusually difficult to sequence by Mass spectrometry, due to the high efficiency of cleavage at the amide bond on the N-terminal of proline residues and the consequently low relative abundance of fragment arising from cleavages at other amide bonds. To fully characterize these proteins by mass spectrometry, specialized approaches to fragmentation are needed for the peptides with high proline content. Our work reported herein focused on the analysis of the set of peptides derived by tryptic cleavage of the salivary protein PRP-3. Two methods of fragmentation were compared: Collision-induced dissociation (CID) and electron capture dissociation (ECD). The data obtained demonstrated that ECD spectra of peptides containing more than 30% proline residues are simpler and easier to interpret than are CID spectra of those peptides. Factors that limit the two methods of fragmentation include the complexity of information contained. in the CID spectra and the low efficiency of ECD processes. A complementary approach using both decomposition methods provides more complete and interpretable sequence information and yielded >93% sequence coverage for the 11-kDa PRP-3 isolated from human saliva.

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