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Titolo:
THE OBSERVATION OF CHAPERONE-LIGAND NONCOVALENT COMPLEXES WITH ELECTROSPRAY-IONIZATION MASS-SPECTROMETRY
Autore:
BRUCE JE; SMITH VF; LIU CL; RANDALL LL; SMITH RD;
Indirizzi:
PACIFIC NW NATL LAB,ENVIRONM & MOL SCI LAB RICHLAND WA 99352 PACIFIC NW NATL LAB,ENVIRONM & MOL SCI LAB RICHLAND WA 99352 WASHINGTON STATE UNIV,DEPT BIOCHEM & BIOPHYS PULLMAN WA 99164
Titolo Testata:
Protein science
fascicolo: 5, volume: 7, anno: 1998,
pagine: 1180 - 1185
SICI:
0961-8368(1998)7:5<1180:TOOCNC>2.0.ZU;2-Y
Fonte:
ISI
Lingua:
ENG
Soggetto:
BINDING PROTEIN; PEPTIDE BINDING; ION-SOURCE; SECB; DNA; DISSOCIATION; BIOMOLECULES; RECOGNITION; PREPROTEIN; SAMPLES;
Keywords:
DISSOCIATION; FOURIER TRANSFORM ION CYCLOTRON RESONANCE; MS/MS; OPPA; PROTEIN EXPORT; SECB; STOICHIOMETRY;
Tipo documento:
Article
Natura:
Periodico
Settore Disciplinare:
Science Citation Index Expanded
Citazioni:
38
Recensione:
Indirizzi per estratti:
Citazione:
J.E. Bruce et al., "THE OBSERVATION OF CHAPERONE-LIGAND NONCOVALENT COMPLEXES WITH ELECTROSPRAY-IONIZATION MASS-SPECTROMETRY", Protein science, 7(5), 1998, pp. 1180-1185

Abstract

Fourier transform ion cyclotron resonance mass spectrometry (FTICR-MS) was applied for the study of noncovalent chaperone SecB-ligand complexes produced in solution and examined in the gas phase with the aid of electrospray ionization (ESI). Since chaperone proteins are believedto recognize and bind only with ligands with nonnative tertiary structure, this work required careful unfolding of the ligand and subsequent reaction with the intact chaperone (the noncovalent tetrameric protein, SecB). A high denaturant concentration was employed to produce nonnative structures of the OppA, and microdialysis of the resulting solutions containing the chaperone-ligand complexes was carried out to rapidly remove the denaturant prior to analysis. Multistage mass spectrometry was essential to the successful study of these complexes since the initial mass spectra indicated extensive adduction that precluded mass measurements, even after microdialysis. However, low energy collisional activation of the ions in the FTICR trap proved useful for adductremoval, and careful control of excitation level preserved the intactcomplexes of interest, revealing a 1:1 SecB:OppA stoichiometry. To our knowledge, these results present the first direct observation of chaperone-ligand noncovalent complexes and the highest molecular weight heterogeneous noncovalent complex observed to date by mass spectrometry. Furthermore, these results highlight the capabilities of FTICR for the study of such complex systems, and the development of a greater understanding of chaperone interactions in protein export.

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Documento generato il 15/07/20 alle ore 14:32:51