Native Ambient Mass Spectrometry Imaging of Ligand-Bound and Metal-Bound Proteins in Rat Brain.

Oliver J Hale,Iain B Styles,Emma K Sisley,Helen J Cooper

doi:10.1021/jacs.1c10032

Oliver J Hale, Iain B Styles + Show 2 more

Open Access

https://doi.org/10.1021/jacs.1c10032

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Abstract

Label-free spatial mapping of the noncovalent interactions of proteins in their tissue environment has the potential to revolutionize life sciences research by providing opportunities for the interrogation of disease progression, drug interactions, and structural and molecular biology more broadly. Here, we demonstrate mass spectrometry imaging of endogenous intact noncovalent protein-ligand complexes in rat brain. The spatial distributions of a range of ligand-bound and metal-bound proteins were mapped in thin tissue sections by use of nanospray-desorption electrospray ionization. Proteins were identified directly from the tissue by top-down mass spectrometry. Three GDP-binding proteins (ADP ribosylation factor ARF3, ARF1, and GTPase Ran) were detected, identified, and imaged in their ligand-bound form. The nature of the ligand was confirmed by multiple rounds of tandem mass spectrometry. In addition, the metal-binding proteins parvalbumin-α and carbonic anhydrase 2 were detected, identified, and imaged in their native form, i.e., parvalbumin-α + 2Ca2+ and carbonic anhydrase + Zn2+. GTPase Ran was detected with both GDP and Mg2+ bound. Several natively monomeric proteins displaying distinct spatial distributions were also identified by top-down mass spectrometry. Protein mass spectrometry imaging was achieved at a spatial resolution of 200 μm.

Highlights

Native ambient mass spectrometry imaging seeks to combine the benefits of native mass spectrometry and ambient mass spectrometry to visualize the spatial distribution of, and obtain structural information on, proteins and their interacting partners
The results demonstrate the mass spectrometry imaging and identification of a range of endogenous ligand-bound and metal-bound protein complexes, in addition to some monomeric proteins with distinct spatial distributions, directly from tissue sections of rat brain at 200 μm spatial resolution
For the protein−ligand complexes, the protein was identified by positive mode top-down mass spectrometry, and the ligand identified by MS3 tandem mass spectrometry using negative mode nano-DESI

Summary

■ INTRODUCTION

Native ambient mass spectrometry imaging seeks to combine the benefits of native mass spectrometry and ambient mass spectrometry to visualize the spatial distribution of, and obtain structural information on, proteins and their interacting partners. We demonstrated native mass spectrometry imaging of folded proteins in thin sections of tissue using the ambient mass spectrometry techniques of both liquid extraction surface analysis (LESA)[1,2] and nanospray desorption electrospray ionization (nano-DESI).[3,4] The present work is the first demonstration of imaging and identification of endogenous protein−ligand and protein− metal (and protein−ligand−metal) complexes directly from tissue. We have applied three approaches for protein identification: top-down nano-DESI of tissue, top-down LESA of tissue, and LESA extraction followed by direct infusion nanoelectrospray top-down mass spectrometry. For some tandem mass spectrometry experiments (see main text), LESA extraction was performed using the Advanced User Interface (AUI) in the Chipsoft software and the sample collected in a microtiter plate before being transferred to custom-made gold coated borosilicate nanoelectrospray emitters.

■ RESULTS AND DISCUSSION

■ CONCLUSION

■ REFERENCES