An organism exposed to a plethora of environmental perturbations undergoes proteomic changes which enable the characterization of total proteins in it. Much of the proteomic information is obtained from genomic data. Additional information on the proteome such as posttranslational modifications, protein–protein interactions, protein localization, metabolic pathways, and so on are deduced using proteomic tools which genomics and transcriptomics fail to offer. The proteomic analysis allows identification of precise changes in proteins, which in turn solve the complexity of microbial population providing insights into the microbial metabolism, cellular pathways, and behavior of microorganisms in new environments. Furthermore, they provide clues for the exploitation of their special features for biotechnological applications. Numerous techniques for the analysis of microbial proteome such as electrophoretic, chromatographic, mass spectrometric-based methods as well as quantitative proteomics are available which facilitate protein separation, expression, identification, and quantification of proteins. An understanding of the potential of each of the proteomic tools has created a significant impact on diverse microbiological aspects and the same has been discussed in this review. © 2020 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim