Analysis of large biomolecules and biomolecular complexes by TOF mass spectrometry

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The proposed research program aims to develop improved mass spectrometric methods for identifying and characterizing large biomolecules and biomolecular complexes. These methods will be applied to various biological problems using time-of-flight (TOF) mass spectrometers of innovative design, constructed in our own laboratory, or in cooperation with our research partner MDS Sciex.      The scientific approach exploits the advantages of TOF instruments for measuring masses, including effectively unlimited m/z range, rapid response, and high sensitivity. Our laboratory has both electrospray and MALDI (matrix-assisted laser desorption/ionization) ion sources coupled to TOF mass spectrometers; the two techniques often provide complementary information about biological molecules.      We plan to continue our extensive program of protein and peptide sequencing, using the hybrid QqTOF mass spectrometer (a cooperative design effort by Sciex and ourselves), which has a new target assembly that enables location of spots on a MALDI target under computer control with much higher speed and accuracy.  Such measurements normally follow the off-line chromatographic separation of complex biological mixtures.      We also plan to couple the recently-discovered electron capture method to a rejuvenated TOF instrument in our laboratory, as described in the accompanying "Research Tools and Instruments" application. This technique for dissociating parent ions tends to cleave peptides along their backbones, leaving the side chains intact, so it is an important complement to the usual collision-induced dissociation, particularly for modified peptides.      A second major activity in our laboratory has been the study of noncovalent complexes. Our ability to investigate these important structures has recently been improved by the construction in our laboratory of an orthogonal injection mass spectrometer specifically designed for the purpose. In particular, this instrument has much higher acceleration voltage than our previous one (16kV vs 4kV), which promises an order of magnitude increase in sensitivity for a typical noncovalent ion.