A Native Mass Spectrometry Systemic View of Cellular Structural Biology

Introduction

Protein structure and consequently function is influenced by the cellular content. The array of splice variants, post-translational modifications, cleavages, and the ability to bind cofactors and drugs is governed by the surrounding cellular milieu. This is determined by internal and external cues such as cell type, developmental stage, stress conditions, disease, and aging, which give rise to an ensemble of distinct protein entities.

Need for New Methods

To study this diversity, there is a need for methods that enable structural studies under “close-to-life” conditions, maintaining the natural environment and biological diversity, features that are often lost during biochemical purifications.

Recent Discoveries

Our recent discovery that native MS can be conducted within crude cellular lysates (direct-MS), while preserving the biological context, offers many new experimental avenues for investigating protein interactions and diversity.

Project Proposal

Here, we propose to take native MS to a new dimension: enabling a systemic view of cellular structural biology by endorsing technological, computational, and methodological developments. To do this we will:

1. Establish a platform for high-throughput screening of protein interactions.
2. Unravel protein interactions in human and other eukaryotic cells.
3. Develop a method for whole-organ direct-MS analysis, unraveling the tissue-specific proteoform landscape.

Biological Systems

Each of our three independent but complementary aims involves a different biological system, going from bacteria through human cells to intact tissues.

Potential Impact

Together, these advances, in conjunction with the high-resolution and sensitivity afforded by current mass spectrometers, have the potential to advance a plethora of fields, including cell biology, pharmacology, and biotechnology. Overall, we anticipate this project will promote the integration of direct-MS into the cellular structural biology toolkit, providing valuable insights not attained by other structural biology methods or artificial intelligence algorithms.