UniDecCD: Deconvolution of Charge Detection-Mass Spectrometry Data

Abstract

Native mass spectrometry (MS) has become a versatile tool for characterizing high-mass complexes and measuring biomolecular interactions. Native MS usually requires the resolution of different charge states produced by electrospray ionization to measure the mass, which is difficult for highly heterogeneous samples that have overlapping and unresolvable charge states. Charge detection-mass spectrometry (CD-MS) seeks to address this challenge by simultaneously measuring the charge and m/z for isolated ions. However, CD-MS often shows uncertainty in the charge measurement that limits the resolution. To overcome this charge state uncertainty, we developed UniDecCD (UCD) software for computational deconvolution of CD-MS data, which significantly improves the resolution of CD-MS data. Here, we describe the UCD algorithm and demonstrate its ability to improve the CD-MS resolution of proteins, megadalton viral capsids, and heterogeneous nanodiscs made from natural lipid extracts. UCD provides a user-friendly interface that will increase the accessibility of CD-MS technology and provide a valuable new computational tool for CD-MS data analysis.

Figure 1.

UCD resolves CD-MS of GroEL similarly to conventional native MS. Conventional native MS of GroEL with A) the raw mass spectrum, B) deconvolved charge vs. m/z, and C) deconvolved mass spectrum with UniDec. CD-MS of GroEL with D) one representative scan of single ions with S/N on the y-axis and the noise level indicated as a red dashed line, E) histogram of charge vs. m/z based on the raw data, and F) mass distribution for the raw CD-MS data, which shows significant uncertainty. In contrast, G) the UCD deconvolved charge vs. m/z and H) the deconvolved CD-MS mass distribution show single charge state resolution.

Figure 2.

UCD deconvolution of CD-MS data from MSP1D1(–) (A–C), BSA (D–F), and ADH (G–I). Each row shows the (A, D, G) raw mass spectrum of one scan of single ions with S/N on the y-axis, (B, E, H) charge vs. m/z histogram after deconvolution, and (C, F, I) deconvolved mass distribution in black overlayed with the raw mass distribution without UCD in green.

Figure 3.

Systems with overlapping charge states measured by CD-MS and deconvolved using UCD. A) AAV viral capsids, B) DPPC nanodiscs, C) E. coli nanodiscs, and D) brain nanodiscs. Each panel (A–D) has a representative scan of single ions (top left), their total deconvolved charge vs. m/z histogram (top right), their deconvolved mass spectrum (bottom) with the raw CD-MS data in green and the UCD deconvolved mass distribution in black. The FWHM values are indicated for raw CD-MS in green and deconvolved in black. The center of the mass and FWHM are listed for a single replicate, and the text indicates the average across three replicate samples.