Poster abstracts

Poster number 44 submitted by Daisy Hamburg

Probing the Topography of Ribonucleoprotein Complexes Using Limited Proteolysis and Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry

Daisy-Malloy Hamburg (Chemistry, University of Cincinnati), Moo-Jin Suh (Chemistry, University of Cincinnati), Patrick A. Limbach (Chemistry, University of Cincinnati)

Abstract:
In the past, limited proteolysis has been employed to attempt to probe the structure of the ribosome with both 2D gel electrophoresis and nuclear magnetic resonance (NMR) as the detection step, with the purpose of determining information about structure of the ribosome in the absence of crystal structures. In this work, limited proteolysis has been combined with matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) allowing for simpler analysis with more reproducible results. In this study, time-dependent digestion was performed on Escherichia coli 30S ribosomal subunits; digestion of the ribosomal proteins allows for the determination of proteins that are resistant to proteolysis. The 30S ribosomal proteins were also analyzed in a variety of condition including their native form to account for the possible resistances to the experimental conditions. MALDI-MS provides accurate mass measurements of the digested proteins, and the resulting data allows for the characterization of surface-exposed regions of the RNP complex. This strategy reveals information about the surface regions more rapidly and from less sample as compared to the prior studies.

Keywords: Ribosome structure, MALDI-MS, Limited Proteolysis