Surface Induced Dissociation: Dissecting Noncovalent Protein Complexes in the Gas phase

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Abstract

The quaternary structures of proteins are both important and of interest to chemists, because many proteins exist as complexes in vivo, and probing these structures allows us to better understand their biological functions. Conventional structural biology methods such as X-ray crystallography and nuclear magnetic resonance provide high-resolution information on the structures of protein complexes and are the gold standards in the field. However, other emerging biophysical methods that only provide low-resolution data (e.g. stoichiometry and subunit connectivity) on the structures of the protein complexes are also becoming more important to scientists. Mass spectrometry is one of these approaches that provide lower than atomic structural resolution, but the approach is higher throughput and provides not only better mass information than other techniques but also stoichiometry and topology. Fragile noncovalent interactions within the protein complexes can be preserved in the gas phase of MS under gentle ionization and transfer conditions. Scientists can measure the masses of the complexes with high confidence to reveal the stoichiometry and composition of the proteins. What makes mass spectrometry an even more powerful method is that researchers can further isolate the protein complexes and activate them in the gas phase to release subunits for more structural information. The caveat is that, upon gas-phase activation, the released subunits need to faithfully reflect the native topology so that useful information on the proteins can be extracted from mass spectrometry experiments. Unfortunately, many proteins tend to favor unfolding upon collision with neutral gas (the most common activation method in mass spectrometers). Therefore, this typically results in limited insights on the quaternary structure of the precursor without further manipulation of other experimental factors. Scientists have observed, however, that valuable structural information can be obtained when the gas-phase proteins are activated by collision with a surface. Subcomplexes released after surface collision are consistent with the native quaternary structure of several protein systems studied, even for a large chaperone protein, GroEL, that approaches megadalton mass. The unique and meaningful data generated from surface induced dissociation (SID) have been attributed to the fast and energetic activation process upon collision with a massive target, the surface. In this Account, we summarize our SID studies of protein complexes, with emphasis on the more recent work on the combination of ion mobility (IM) with SID. IM has gained popularity over the years not only as a gas-phase separation technique but also as a technique with the ability to measure the size and shape of the proteins in the gas phase. Incorporation of IM before SID allows different conformations of a protein to be separated and examined individually by SID for structural details. When IM is after SID, the cross sections of the SID products can be measured, providing insight on the dissociation pathways, which may mimic disassembly pathways. Furthermore, the separation by IM greatly reduces the peak overlapping (same m/z) and coalescence (merging) of SID products, improving the resolving power of the method. While there are still many unanswered questions on the fundamental properties of gas-phase proteins and a need for further research, our work has shown that SID can be a complementary gas-phase tool providing useful information for studying quaternary structures of noncovalent protein complexes.

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