Attomole protein characterization by capillary electrophoresis-mass spectrometry.
Electrospray ionization with an ultralow flow rate (</=4 nanoliters per minute) was used to directly couple capillary electrophoresis with tandem mass spectrometry for the analysis and identification of biomolecules in mixtures. A Fourier transform mass spectrometer provided full spectra (>30 kilodaltons) at a resolving power of approximately 60,000 for injections of 0.7 x 10(-18) to 3 x 10(-18) mole of 8- to 29-kilodalton proteins with errors of <1 dalton in molecular mass. Using a crude isolate from human blood, a value of 28,780.6 daltons (calculated, 28,780.4 daltons) was measured for carbonic anhydrase, representing 1 percent by weight of the protein in a single red blood cell. Dissociation of molecular ions from 9 x 10(-18) mole of carbonic anhydrase gave nine sequence-specific fragment ions, more data than required for unique retrieval of this enzyme from the protein database.
Valaskovic
G A
GA
Department of Chemistry, Baker Laboratory, Cornell University, Ithaca, NY 14853, USA.
Kelleher
N L
NL
McLafferty
F W
FW
D000818:Animals; D002256:Carbonic Anhydrases / Q000032:analysis* / Q000737:chemistry; D002417:Cattle; D003574:Cytochrome c Group / Q000032:analysis / Q000737:chemistry; D019075:Electrophoresis, Capillary / Q000379:methods*; D006736:Horses; D006801:Humans; D013058:Mass Spectrometry / Q000379:methods*; D008970:Molecular Weight; D011506:Proteins / Q000032:analysis* / Q000737:chemistry; D012680:Sensitivity and Specificity; D017550:Spectroscopy, Fourier Transform Infrared; D014452:Ubiquitins / Q000032:analysis / Q000737:chemistry
D003574:Cytochrome c Group; D011506:Proteins; D014452:Ubiquitins; D002256:Carbonic Anhydrases