Quantitative proteomics in the field of microbiology

Abstract

Quantitative proteomics has become an indispensable analytical tool for microbial research. Modern microbial proteomics covers a wide range of topics in basic and applied research from in vitro characterization of single organisms to unravel the physiological implications of stress/starvation to description of the proteome content of a cell at a given time. With the techniques available, ranging from classical gel-based procedures to modern MS-based quantitative techniques, including metabolic and chemical labeling, as well as label-free techniques, quantitative proteomics is today highly successful in sophisticated settings of high complexity such as host-pathogen interactions, mixed microbial communities, and microbial metaproteomics. In this review, we will focus on the vast range of techniques practically applied in current research with an introduction of the workflows used for quantitative comparisons, a description of the advantages/disadvantages of the various methods, reference to hallmark publications and presentation of applications in current microbial research.

Life Stage-specific Proteomes of Legionella pneumophila Reveal a Highly Differential Abundance of Virulence-associated Dot/Icm effectors

“…Biotinylation of proteins was performed by incubation of intact cells with Sulfo-NHS-SS biotin, which is advertised as membrane impermeable and therefore is expected to react specifically with proteins residing in the membrane or being exposed to the exterior (175,176). By means of that method coupled to subsequent enrichment analysis, we detected a variety of known and potentially surface-exposed proteins as well as outer membrane-interacting proteins, such as Mip, EnhA, TolB, OmpH/Lpg0507, and Ttg2D/Lpg0844 (supplemental Table S6) (75,91,177). However, some studies also report on the labeling of intracellular proteins when using Sulfo-NHS-SS biotin (178).…”

Impact of the omic technologies for understanding the modes of action of biological control agents against plant pathogens

“…chitinases, cellulases and proteases) (Aegerter and Gordon 2006). For the 'discovery driven' approach, several proteomic methodologies are available, they can be distinguished as the gel-based and the gel-free methods and excellent reviews recently discussed technological advances in the microbial proteomics (Armengaud 2013;Gil and Monteoliva 2014;Otto et al 2014Otto et al , 2012Oudenhove and Devreese 2013). Briefly, the gel-based approaches are based on protein separation by the two-dimensional (2D) gel electrophoresis followed by protein spot isolation and identification by mass spectrometry, while the gel-free proteomics is based on the enzymatic digestion of the protein mixture followed by liquid chromatography and identification of peptide sequences by mass spectrometry (Otto et al 2012;Wöhlbrand et al 2013).…”

“…Progress has been made in the field of microbial proteomics thanks to developments in sample preparation, high resolution separation techniques, high performance mass spectrometers and software for protein identification and quantification (Armengaud 2013;Otto et al 2014Otto et al , 2012Oudenhove and Devreese 2013). However, there still exist different technical challenges and limitations since protein separation and analysis are inherently skill-based and are difficult to automate.…”

Application of proteomics in non-small-cell lung cancer

“…Incorporating with powerful statistical software, this high-throughput technology enables the identification and quantitation of a great coverage of peptides and proteins with high sensitivity. [3] MS-based proteomics is getting mature in the last decade and is now combining with other techniques to enhance reproducibility and accuracy. [4] Apart from label-free quantification, the stable isotope labeling method has risen as a commonly used approach for proteomics study.…”