Abstract
Chemical cross-linking of proteins is a well-established method for structural mapping of small protein complexes. When combined with mass spectrometry, cross-linking can reveal protein topology and identify contact sites between the peptide surfaces. When applied to surface-exposed proteins from pathogenic organisms, the method can reveal structural details that are useful in vaccine design. In order to investigate the possibilities of applying cross-linking on larger protein complexes, we selected the urease enzyme from Helicobacter pylori as a model. This membrane-associated protein complex consists of two subunits: α (26.5 kDa) and β (61.7 kDa). Three (αβ) heterodimers form a trimeric (αβ)3 assembly which further associates into a unique dodecameric 1.1 MDa complex composed of four (αβ)3 units. Cross-linked peptides from trypsin-digested urease complex were analyzed by Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) and molecular modeling. Two potential cross-linked peptides (present in the cross-linked sample but undetectable in α, β, and native complex) were assigned. Molecular modeling of urease αβ complex and trimeric urease units (αβ)3 revealed a linkage site between the α-subunit and the β-subunit, and an internal cross-linkage in the β-subunit.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 137-144 |
| Number of pages | 8 |
| Journal | International Journal of Mass Spectrometry |
| Volume | 234 |
| Issue number | 1-3 |
| DOIs | |
| State | Published - May 1 2004 |
| Externally published | Yes |
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Keywords
- Cross-linking
- FT-ICR MS
- FTMS
- Molecular modeling
- Urease
ASJC Scopus subject areas
- Physical and Theoretical Chemistry
- Spectroscopy
Cite this
Chemical cross-linking of the urease complex from Helicobacter pylori and analysis by Fourier transform ion cyclotron resonance mass spectrometry and molecular modeling. / Carlsohn, Elisabet; Ångström, Jonas; Emmett, Mark; Marshall, Alan G.; Nilsson, Carol L.
In: International Journal of Mass Spectrometry, Vol. 234, No. 1-3, 01.05.2004, p. 137-144.Research output: Contribution to journal › Article
}
TY - JOUR
T1 - Chemical cross-linking of the urease complex from Helicobacter pylori and analysis by Fourier transform ion cyclotron resonance mass spectrometry and molecular modeling
AU - Carlsohn, Elisabet
AU - Ångström, Jonas
AU - Emmett, Mark
AU - Marshall, Alan G.
AU - Nilsson, Carol L.
PY - 2004/5/1
Y1 - 2004/5/1
N2 - Chemical cross-linking of proteins is a well-established method for structural mapping of small protein complexes. When combined with mass spectrometry, cross-linking can reveal protein topology and identify contact sites between the peptide surfaces. When applied to surface-exposed proteins from pathogenic organisms, the method can reveal structural details that are useful in vaccine design. In order to investigate the possibilities of applying cross-linking on larger protein complexes, we selected the urease enzyme from Helicobacter pylori as a model. This membrane-associated protein complex consists of two subunits: α (26.5 kDa) and β (61.7 kDa). Three (αβ) heterodimers form a trimeric (αβ)3 assembly which further associates into a unique dodecameric 1.1 MDa complex composed of four (αβ)3 units. Cross-linked peptides from trypsin-digested urease complex were analyzed by Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) and molecular modeling. Two potential cross-linked peptides (present in the cross-linked sample but undetectable in α, β, and native complex) were assigned. Molecular modeling of urease αβ complex and trimeric urease units (αβ)3 revealed a linkage site between the α-subunit and the β-subunit, and an internal cross-linkage in the β-subunit.
AB - Chemical cross-linking of proteins is a well-established method for structural mapping of small protein complexes. When combined with mass spectrometry, cross-linking can reveal protein topology and identify contact sites between the peptide surfaces. When applied to surface-exposed proteins from pathogenic organisms, the method can reveal structural details that are useful in vaccine design. In order to investigate the possibilities of applying cross-linking on larger protein complexes, we selected the urease enzyme from Helicobacter pylori as a model. This membrane-associated protein complex consists of two subunits: α (26.5 kDa) and β (61.7 kDa). Three (αβ) heterodimers form a trimeric (αβ)3 assembly which further associates into a unique dodecameric 1.1 MDa complex composed of four (αβ)3 units. Cross-linked peptides from trypsin-digested urease complex were analyzed by Fourier transform ion cyclotron resonance mass spectrometry (FT-ICR MS) and molecular modeling. Two potential cross-linked peptides (present in the cross-linked sample but undetectable in α, β, and native complex) were assigned. Molecular modeling of urease αβ complex and trimeric urease units (αβ)3 revealed a linkage site between the α-subunit and the β-subunit, and an internal cross-linkage in the β-subunit.
KW - Cross-linking
KW - FT-ICR MS
KW - FTMS
KW - Molecular modeling
KW - Urease
UR - http://www.scopus.com/inward/record.url?scp=2442546545&partnerID=8YFLogxK
UR - http://www.scopus.com/inward/citedby.url?scp=2442546545&partnerID=8YFLogxK
U2 - 10.1016/j.ijms.2004.02.021
DO - 10.1016/j.ijms.2004.02.021
M3 - Article
AN - SCOPUS:2442546545
VL - 234
SP - 137
EP - 144
JO - International Journal of Mass Spectrometry
JF - International Journal of Mass Spectrometry
SN - 1387-3806
IS - 1-3
ER -