Abstract
Cross-linking analysis of protein complexes and structures by tandem mass spectrometry (MS/MS) has advantages in speed, sensitivity, specificity, and the capability of handling complicated protein assemblies. However, detection and accurate assignment of the cross-linked peptides are often challenging due to their low abundance and complicated fragmentation behavior in collision-induced dissociation (CID). To simplify the MS analysis and improve the signal-to-noise ratio of the cross-linked peptides, we developed a novel peptide enrichment strategy that utilizes a cross-linker with a cryptic thiol group and using beads modified with a photocleavable cross-linker. The functional cross-linkers were designed to react with the primary amino groups in proteins. Human serum albumin was used as a model protein to detect intra- and intermolecular cross-linkages. Use of this protein-free selective retrieval method eliminates the contamination that can result from avidin - biotin based retrieval systems and simplifies data analysis. These features may make the method suitable to investigate protein - protein interactions in biological samples.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 7149-7159 |
| Number of pages | 11 |
| Journal | Analytical Chemistry |
| Volume | 81 |
| Issue number | 17 |
| DOIs | |
| State | Published - Sep 1 2009 |
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ASJC Scopus subject areas
- Analytical Chemistry
Cite this
Nonprotein based enrichment method to analyze peptide cross-linking in protein complexes. / Yan, Funing; Che, Fa Yun; Rykunov, Dmitry; Nieves, Edward; Fiser, Andras; Weiss, Louis M.; Angeletti, Ruth Hogue.
In: Analytical Chemistry, Vol. 81, No. 17, 01.09.2009, p. 7149-7159.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Nonprotein based enrichment method to analyze peptide cross-linking in protein complexes
AU - Yan, Funing
AU - Che, Fa Yun
AU - Rykunov, Dmitry
AU - Nieves, Edward
AU - Fiser, Andras
AU - Weiss, Louis M.
AU - Angeletti, Ruth Hogue
PY - 2009/9/1
Y1 - 2009/9/1
N2 - Cross-linking analysis of protein complexes and structures by tandem mass spectrometry (MS/MS) has advantages in speed, sensitivity, specificity, and the capability of handling complicated protein assemblies. However, detection and accurate assignment of the cross-linked peptides are often challenging due to their low abundance and complicated fragmentation behavior in collision-induced dissociation (CID). To simplify the MS analysis and improve the signal-to-noise ratio of the cross-linked peptides, we developed a novel peptide enrichment strategy that utilizes a cross-linker with a cryptic thiol group and using beads modified with a photocleavable cross-linker. The functional cross-linkers were designed to react with the primary amino groups in proteins. Human serum albumin was used as a model protein to detect intra- and intermolecular cross-linkages. Use of this protein-free selective retrieval method eliminates the contamination that can result from avidin - biotin based retrieval systems and simplifies data analysis. These features may make the method suitable to investigate protein - protein interactions in biological samples.
AB - Cross-linking analysis of protein complexes and structures by tandem mass spectrometry (MS/MS) has advantages in speed, sensitivity, specificity, and the capability of handling complicated protein assemblies. However, detection and accurate assignment of the cross-linked peptides are often challenging due to their low abundance and complicated fragmentation behavior in collision-induced dissociation (CID). To simplify the MS analysis and improve the signal-to-noise ratio of the cross-linked peptides, we developed a novel peptide enrichment strategy that utilizes a cross-linker with a cryptic thiol group and using beads modified with a photocleavable cross-linker. The functional cross-linkers were designed to react with the primary amino groups in proteins. Human serum albumin was used as a model protein to detect intra- and intermolecular cross-linkages. Use of this protein-free selective retrieval method eliminates the contamination that can result from avidin - biotin based retrieval systems and simplifies data analysis. These features may make the method suitable to investigate protein - protein interactions in biological samples.
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U2 - 10.1021/ac900360b
DO - 10.1021/ac900360b
M3 - Article
VL - 81
SP - 7149
EP - 7159
JO - Analytical Chemistry
JF - Analytical Chemistry
SN - 0003-2700
IS - 17
ER -