High Definition Mass Spectrometry as a Tool for Structural Investigation of High m/z Ion Species

Here we present the use of high-efficiency IMS (Triwave technology) on a SYNAPT High Definition Mass Spectrometry (HDMS) System for analysis of high m/z caesium iodide clusters over the m/z range 1,000–20,000. This demonstrates the utility of the SYNAPT HDMS System for the mass measurement of high m/z species, such as in the analysis of non-covalent protein complexes.

Over the past 10 years, interest in high mass non-covalent analysis has increased due to the ability of the current mass spectrometers and electrospray sources to preserve the non-covalent interactions, allowing one to analyze compounds in their native conformation and stoichiometry.¹ The transfer of non-covalently associated complexes from solution to the gas phase using electrospray ionization generally results in the formation of ions possessing relatively few charges. As a result, these species appear high on the m/z scale, making Time-of-Flight mass spectrometry ideal for their mass analysis. The utility of ion mobility spectrometry (IMS) in probing the structures of relatively large complexes has been highlighted previously.¹

Here we present the use of high-efficiency IMS (Triwave technology) on a SYNAPT High Definition Mass Spectrometry (HDMS) System for analysis of high m/z caesium iodide clusters over the m/z range 1,000–20,000. This demonstrates the utility of the SYNAPT HDMS System for the mass measurement of high m/z species, such as in the analysis of non-covalent protein complexes.

The instrument used in this study was a SYNAPT HDMS System, which combines high-efficiency ion mobility based measurements and separations with a hybrid quadrupole orthogonal acceleration Time-of-Flight (oa-ToF) mass spectrometer, Figure 1.² Samples were introduced with a borosilcate-glass nano electrospray-spray tip and sampled into the vacuum system. The ions pass through a quadrupole mass filter to the enabling Triwave device. Triwave consists of three travelling wave (T-Wave) ion guides. The TRAP T-Wave accumulates ions (with high efficiency), after which these ions are released as discrete packets into the IMS T-Wave, where the ion mobility separation of ions is performed. The TRANSFER T-Wave is used to deliver the ion mobility-separated ions into the oa-ToF analyzer. Each IMS separation was 51 ms long and the ions were released from the TRAP T-wave in 500 μs wide packets. The gas pressure in the TRAP and TRANSFER T-Wave regions was 0.07 mbar (Argon) and the pressure in the IMS T-Wave was 0.5 mbar (Nitrogen). The traveling wave used in the IMS T-Wave for ion mobility separation was operated at a velocity of 250 m/sec. The wave amplitude was ramped from 0 to 30 V over the period of the mobility separation for optimum performance over the large m/z range used (m/z 1,000 to 32,000).

Upon MS acquisition of a concentrated caesium iodide solution, intense ion clusters can be observed as high as m/z 20,000, with each cluster’s composition based on the formula Cs(n+1)In. From the mass spectrum generated it is evident that a number of overlapping series, differing in charge state and intensity profile, are present over the m/z scale (Figure 2). However, the HDMS (IMS/MS) analysis clearly illustrates discrete distributions, which are related by their m/z and drift-time, as shown in Figure 3.

The drift-times of the cluster ions seem to increase monotonically with increasing m/z values for the different charge state species, although with increasing charge state series up to +4, distinct m/z stability regions become clear, as shown in Figure 4.

• HDMS provides greater definition of even relatively simple samples, such as inorganic CsI clusters
  
• High mass analysis of CsI clusters up to m/z 20,000 has been shown
  
• CsI clusters have been successfully separated into five charge states series, demonstrating the ability of HDMS (high-efficiency IMS combined with oa-ToF mass spectrometry) for differentiating high mw species based on their mobility
  
• CsI can be used to tune, optimize, and calibrate the SYNAPT HDMS System over a wide m/z range prior to high mass acquisitions being carried out

1. Ruotolo, Giles, Campuzano, Sandercock, Bateman & Robinson. Science. 310 (2005) 1658–1661.
   
2. Pringle, Giles, Wildgoose, Williams, Slade, Thalassinos, Bateman, Bowers, Scrivens. Int. J. Mass Spectrom., 261 (2007) 1–12.