Multipole-storage-assisted dissociation for the characterization of large proteins and simple protein mixtures by ESI-FTICR-MS.
In Fourier transform ion cyclotron resonance mass spectrometry, collisionally activated dissociation (CAD) typically is accomplished within the analyzer ion cell. An alternative approach of multipole-storage-assisted dissociation (MSAD) has previously been demonstrated by inducing collisional fragmentation in the external multipole that is usually employed for ion accumulation. To explore the utility of MSAD for interrogating intact proteins and simple protein mixtures in a multiplexed manner, we have investigated the means of controlling the collisional energy and the fragmentation pattern for this experimental approach. With protein samples in the low micromolar concentration range, the two major experimental parameters affecting MSAD in the hexapole region were found to be the dc offset voltage and accumulation time. While low-energy MSAD of intact proteins yields fragment ions similar to sustained off resonance irradiation collision-activated dissociation (SORI-CAD), high-energy MSAD induces sequential fragmentation for intact proteins to yield a rich variety of singly charged ions in the m/z 600-1200 Da region. Each of the seven proteins (Mr range of 8.5-116 kDa) examined in this study exhibited their own characteristic MSAD fragmentation pattern, which could be used as a signature of the presence of a given protein, even in a mixture. In addition, any MSAD fragment can be isolated and dissociated further by SORI-CAD in an MS3-type experiment inside the FTICR analyzer cell. This presents a novel way to interrogate the identities of these fragment ions as well as obtain amino acid sequence tag information that can be used to identify proteins from mixtures.
Pan
Chongle
C
Genome Science and Technology Graduate School, Oak Ridge National Laboratory-University of Tennessee, Oak Ridge, Tennessee 37831-6131, USA.
Hettich
Robert L
RL
D000595:Amino Acid Sequence; D001059:Apoproteins / Q000032:analysis / Q000737:chemistry; D007782:Lactoglobulins / Q000032:analysis / Q000737:chemistry; D008970:Molecular Weight; D009211:Myoglobin / Q000032:analysis / Q000737:chemistry; D011506:Proteins / Q000032:analysis* / Q000737:chemistry*; D012709:Serum Albumin / Q000032:analysis / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D017550:Spectroscopy, Fourier Transform Infrared / Q000379:methods*; D025801:Ubiquitin / Q000032:analysis / Q000737:chemistry
D001059:Apoproteins; D007782:Lactoglobulins; D009211:Myoglobin; D011506:Proteins; D012709:Serum Albumin; D025801:Ubiquitin; C010857:apomyoglobin