Protein identification via surface-induced dissociation in an FT-ICR mass spectrometer and a patchwork sequencing approach.
Surface-induced dissociation (SID) and collision-induced dissociation (CID) are ion activation techniques based on energetic collisions with a surface or gas molecule, respectively. One noticeable difference between CID and SID is that SID does not require a collision gas for ion activation and is, therefore, directly compatible with the high vacuum requirement of Fourier transform ion cyclotron resonance (FT-ICR) mass spectrometers. Eliminating the introduction of collision gas into the ICR cell for collisional activation dramatically shortens the acquisition time for MS/MS experiments, suggesting that SID could be utilized for high-throughput MS/MS studies in FT-ICR MS. We demonstrate for the first time the utility of SID combined with FT-ICR MS for protein identification. Tryptic digests of standard proteins were analyzed using a hybrid 6-tesla FT-ICR mass spectrometer with SID and CID capabilities. SID spectra of mass-selected singly and doubly charged peptides were obtained using a diamond-coated target mounted at the rear trapping plate of the ICR cell. The broad internal energy distribution deposited into the precursor ion following collision with the diamond surface allowed a variety of fragmentation channels to be accessed by SID. Composition and sequence qualifiers produced by SID of tryptic peptides were used to improve the statistical significance of database searches. Protein identification MASCOT scores obtained using SID were comparable or better than scores obtained using sustained off-resonance irradiation collision-induced dissociation (SORI-CID), the conventional ion activation technique in FT-ICR MS.
School of Chemistry and Biochemistry, Georgia Institute of Technology, Atlanta, Georgia, USA.
Facundo M
FM
Fernandez
Vicki H
VH
Wysocki
Jean H
JH
Futrell
Julia
J
Laskin
D000199:Actins / Q000032:analysis; D000426:Alcohol Dehydrogenase / Q000032:analysis; D000595:Amino Acid Sequence; D000818:Animals; D045304:Cytochromes c / Q000032:analysis; D006736:Horses; D008969:Molecular Sequence Data; D018482:Muscle, Skeletal / Q000737:chemistry; D011506:Proteins / Q000032:analysis*; D011817:Rabbits; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods*; D015003:Yeasts / Q000201:enzymology
D000199:Actins; D011506:Proteins; D045304:Cytochromes c; D000426:Alcohol Dehydrogenase