Mass spectrometry and many kinds of related techniques have made a big contribution to proteomic research. On the other hand, various types of approaches other than mass spectrometry have also been used in proteomic analyses, making the best use of their characters and advantages. Large-scale data are vital for efficient finding of molecular functions and mechanisms in life. In addition, it is also significant to understand living organism as a system where whole molecules, such as proteome, coordinate or compete with one other to drive biological activity. In this review, I will introduce a variety of proteomic approaches independent of mass spectrometry, especially for budding yeast, and describe my perspective about what is required to further expand the proteomic research.
Many proteins in the organisms function by interacting with other proteins to form complexes. Especially in higher organisms such as human, the formation of complex protein is one of the mechanisms that regulate protein function. Therefore, the analysis of interacting proteins is an important issue to understand the complicated regulatory mechanisms that sustain the higher organisms. As a method to analyze interacting proteins, the BioID technique has been developed in which a proximity biotinylation enzyme is fused to the target protein for labeling the proximal proteins with biotin. Recently, we have succeeded in developing a novel proximity biotinylation enzyme, AirID, by using an algorithm to design an ancestral protein in silico based on huge genome sequence data. In this review, I will introduce an overview of BioID, TurboID and AirID, which are representative proximity biotinylation enzymes, and examples of protein interaction analysis using AirID mainly.
Metaproteomics using mass spectrometry (MS) is a powerful method for classifying the very complex microorganisms in samples such as marine water, soil, feces, and saliva, and for profiling all the proteins expressed by all the microorganisms. Metagenomics can classify and profile the genes of the microorganisms in a sample, while metaproteomics can profile the proteins actually expressed to gain functional knowledge and understand what the microorganisms are doing. The analysis must be on a large scale, since the number of bacteria in the human intestine is larger than 1,000 species and 39 trillion, and the variety of species in each individual is very large. In this review, we discuss recent trends in metaproteomics of intestinal bacteria, including analysis software, protein sequence databases for intestinal bacteria, and issues related to large-scale analysis.