Proteome-wide cellular protein concentrations of the human pathogen Leptospira interrogans

Abstract

Mass-spectrometry-based methods for relative proteome quantification have broadly affected life science research. However, important research directions, particularly those involving mathematical modelling and simulation of biological processes, also critically depend on absolutely quantitative data--that is, knowledge of the concentration of the expressed proteins as a function of cellular state. Until now, absolute protein concentration measurements of a considerable fraction of the proteome (73%) have only been derived from genetically altered Saccharomyces cerevisiae cells, a technique that is not directly portable from yeast to other species. Here we present a mass-spectrometry-based strategy to determine the absolute quantity, that is, the average number of protein copies per cell in a cell population, for a large fraction of the proteome in genetically unperturbed cells. Applying the technology to the human pathogen Leptospira interrogans, a spirochete responsible for leptospirosis, we generated an absolute protein abundance scale for 83% of the mass-spectrometry-detectable proteome, from cells at different states. Taking advantage of the unique cellular dimensions of L. interrogans, we used cryo-electron tomography morphological measurements to verify, at the single-cell level, the average absolute abundance values of selected proteins determined by mass spectrometry on a population of cells. Because the strategy is relatively fast and applicable to any cell type, we expect that it will become a cornerstone of quantitative biology and systems biology.

Figure 1. Large scale determination of cellular protein concentrations

A) Natural logarithm of extracted precursor ion intensities plotted against the natural logarithm of copies/cell for 16 proteins quantified by SRM. B) Distribution of error rates determined by bootstrapping. C) Slice through tomographic reconstruction, substructures are marked color-coded as described below (scale bar 200 nm). The inset shows a close-up of methyl-accepting proteins (scale bar 100 nm). The boxes display the gene products making up the different components of methyl-accepting proteins (green), periplasmic flagella (dark blue), the flagellar stator (transparent red) and rotor (dark red).

Figure 2. Abundance levels of selected protein groups (by GeneOntology)

The number of genes, number of identified proteins and copies per cell for two cellular states (control and ciprofloxacin treated). Please note that hypothetical proteins are underrepresented in the copies per cell calculation as compared to gene numbers, while members of the protein folding, encapsulating structure and electron transport group are largely overrepresented.