News and Publications

Characterizing Biomolecular Interactions

G. Siuzdak, J. Boydston, B. Bothner, T. Hollenbeck, J. Wu, K.S. Chatman, K. Harris, A. Aparicio, R. North, D. Owens, X. Gao, J.K. Lewis

In the late 1980s, mass spectrometry emerged as an important tool in biomolecular research, and although the usefulness of this method has increased significantly in the past decade, its full potential is far from being realized. The value of mass spectrometry lies in the unique ability to accurately characterize the structure and dynamics of biomolecules by using only femtomoles to picomoles of material.

Our interests focus on understanding the functions of biomolecules and on developing novel methods for examining the interactions and reactions of these complexes. For instance, the goal in our studies of protein structure is to determine specific sites of protein interactions and activity, with special emphasis on viral proteins and enzymes. Our unique application of mass spectrometry to viral analysis allows us to examine both local and global viral structure, thus gaining interesting insights into the structure of proteins on the viral surface.

Despite the detailed structural models created for viruses, little is known about the mechanisms used by the capsid proteins that facilitate binding to host cells, penetration of the cell membrane, and release of nucleic acid. We have used matrix-assisted laser desorption/ionization mass spectrometry combined with limited proteolysis to examine the viral capsids of human rhinovirus and flock house virus. Using only microgram quantities of virus, we obtained novel information about the dynamic nature of the viral surface (Fig. 1) and about the interaction of human rhinovirus with antiviral agents. Moreover, we have been able to characterize posttranslational modifications in human rhinovirus and have used high-resolution Fourier transform mass spectrometry to detect viral mutants (Fig. 2).

We are also aware of the power of automation in combination with mass spectrometry. To this end, we are developing assays that can be used for virtually any chemical or biological reaction. For instance, with the development of a novel mass spectrometry assay that is both automated and quantitative, new enzyme inhibitors and antiviral agents can be detected without the use of a chromophore or radiolabeling. In addition, this method allows monitoring of more than 1000 reactions in a single day. Clearly, the usefulness of mass spectrometry extends far beyond molecular weight characterization and offers great prospects for understanding dynamic biomolecular interactions. More information on mass spectrometry in biochemical research is available at http://masspec.scripps.edu/.

PUBLICATIONS

Bothner, B., Dong, X.F., Bibbs, L., Johnson, J.E., Siuzdak, G. Evidence of viral capsid dynamics using limited proteolysis and mass spectrometry. J. Biol. Chem. 273:673, 1998.

Lewis, J.K., Bendahmane, M., Smith, T.J., Beachy, R.N., Siuzdak, G. Identification of viral mutants by mass spectrometry. Proc. Natl. Acad. Sci. U.S.A., in press.

Lewis, J.K., Bothner, B., Smith, T.J., Siuzdak, G. Antiviral agent blocks breathing of common cold virus. Proc. Natl. Acad. Sci. U.S.A., in press.

Owens, D.R., Bothner, B., Harris, K., Phung, Q., Siuzdak, G. Aspects of oligonucleotide and peptide sequencing with mass spectrometry. Bioorg. Med. Chem., in press.

Siuzdak, G. Mass spectrometry. In: The Encyclopedia of Molecular Biology. Creighton, T.E. (Ed.). Wiley, New York, in press.

Siuzdak, G. Probing molecular diversity with mass spectrometry. J. Assoc. Lab. Automation, in press.

Siuzdak, G. Probing viruses with mass spectrometry. J. Mass Spectrom. 33:203, 1998.

Siuzdak, G., Henriksen, S. New directions in the analysis of brain substances related to sleep and wakefulness. In: Molecular Regulation of Arousal States. Lydic, R. (Ed.). CRC Press, Boca Raton, FL, 1997, p. 181.

Siuzdak, G., Lewis, J.K. Applications of mass spectrometry in combinatorial chemistry. Combin. Chem., in press.