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

Abstract

Mass spectrometry based methods for relative proteome quantification have broadly impacted life science research. However, important research directions, particularly those involving mathematical modeling and simulation of biological processes, also critically depend on absolutely quantitative data, i.e. knowledge of the concentration of the expressed proteins as a function of cellular state. Until now, absolute protein concentration measurements of a significant fraction of the proteome (73%) have only been derived from genetically altered S. cerevisiae cells 1, a technique that is not directly portable from yeast to other species. In this study we developed and applied a mass spectrometry based strategy to determine the absolute quantity i.e. the average number of protein copies per cell in a cell population, for a significant fraction of the proteome in genetically unperturbed cells. Applying the technology to the human pathogen Leptospira interrogans, a spirochete responsible for Leptospirosis 4, 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 (cryoET) 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. As 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.

Global quantification of mammalian gene expression control

“…S6 B). iBAQ values correlated well with known absolute protein amounts over at least four orders of magnitude and had a higher precision and accuracy than alternative measures of absolute protein abundance (data not shown) 23,24 .…”

“…Absolute protein copy numbers were inferred from mass spec data 23,24 . To this end, we used the sum of peak intensities of all peptides matching to a specific protein.…”

High-coverage proteome analysis reveals the first insight of protein modification systems in the pathogenic spirochete Leptospira interrogans

“…This represents the second highest proteome coverage so far for prokaryotes, only lower than that (88.6%) of the simple species Mycoplasma mobile (~0.78 Mb), which has a genome of only a onesixth size of that of L. interrogans [30]. Recently, 2 221 (60.7%) out of 3 658 predicted proteins in L. interrogans serovar Copenhageni were identified [14], among which 2 178 had orthologs in the serovar Lai and 1 982 of these orthologs were identified in our study.…”

“…Meanwhile, a wide range of hypothetical proteins were identified to be expressed in the serovar Lai. In addition, we compared protein identification in the serovar Lai with that in the serovar Copenhageni [14] and found that these two serovars had similar protein expression profiles in vitro: 89.0% expressed proteins in the serovar Copenhageni were also identified in the serovar Lai.…”

“…The genome sequences of the pathogenic L. interrogans serovar Lai and serovar Copenhageni, the pathogenic L. borgpetersenii serovar Hardjo and the saprophytic L. biflexa serovar Patoc have been reported [3][4][5][6]. The accomplishment of these sequencing projects greatly facilitated the studies of Leptospira physiology and pathology at the genomic [4][5][6], transcriptomic [7,8] and proteomic levels [9][10][11][12][13][14]. However, there are discrepancies in the annotations for the same genome sequences by different institutions or by using diverse prediction methodologies, which hinders further studies [8].…”