Multi-Omic Single-Shot Technology for Integrated Proteome and Lipidome Analysis.

Yuchen He,Evgenia Shishkova,Edrees H Rashan,Vanessa Linke,David J Pagliarini,Michael S Westphall,Joshua J Coon,Adam Jochem,Alexander S Hebert,Katherine A Overmyer

doi:10.1021/acs.analchem.0c04764

Abstract

Mass spectrometry (MS) serves as the centerpiece technology for proteome, lipidome, and metabolome analysis. To gain a better understanding of the multifaceted networks of myriad regulatory layers in complex organisms, integration of different multiomic layers is increasingly performed, including joint extraction methods of diverse biomolecular classes and comprehensive data analyses of different omics. Despite the versatility of MS systems, fractured methodology drives nearly all MS laboratories to specialize in analysis of a single ome at the exclusion of the others. Although liquid chromatography-mass spectrometry (LC-MS) analysis is similar for different biomolecular classes, the integration on the instrument level is lagging behind. The recent advancements in high flow proteomics enable us to take a first step towards integration of protein and lipid analysis. Here, we describe a technology to achieve broad and deep coverage of multiple molecular classes simultaneously through multi-omic single-shot technology (MOST), requiring only one column, one LC-MS instrument, and a simplified workflow. MOST achieved great robustness and reproducibility. Its application to a Saccharomyces cerevisiae study consisting of 20 conditions revealed 2842 protein groups and 325 lipids and potential molecular relationships.

Highlights

To test our hypothesis that peptides and lipids can be co-analyzed in a single LC-Mass spectrometry (MS) methodology, we first sequentially loaded a complex mixture of lipids and peptides from a yeast cell lysate onto an RP LC column
Its addition led to a slight reduction in peptide identifications but a substantial increase in lipid identifications compared to no-ammonium analyses: 5.8% fewer unique peptides and 3.3% fewer protein groups as compared to an 11.6% gain in lipid identifications
Comparable results were obtained for mass error (0.40 vs. 0.41 absolute median ppm for peptides and 2.73 vs. 2.72 absolute median ppm for lipids) and identifications (< 10 identified proteins or lipids differed between single-ome high flow platforms and our newly developed multi-omic single-shot technology (MOST) platform) (Supplementary Figure 2c-e)

Summary

Introduction

We typically detect ~ 150 lipids, a number comparable to the amount observed when we perform lipidomics separately on yeast (Supplementary Figure 1c). The BEH column led to more identifications in both proteomic and lipidomic analyses: 1.3% more unique peptides and proteins, 14.9% more lipids (Supplementary Figure 1f-g).

Results

Conclusion