Table 4.

Proteomic and secretomic tools used in secondary metabolite study

Tool and techniques	Mechanism	Applications	References
2-D PAGE and Isoelectric focusing (IEF)	Separation and identification of proteins by their charge-to-mass ratio	Helps in the identification of posttranslational modified natural protein and SM	Shankar et al. (2019)
MALDI-ToF/ToF	Ionize biological molecule to identify their mass and sequences	Ionized biological sample gives all about the molecular weight, protein sequence, and three-dimensional structure of metabolites	Shankar et al. (2019)
Electrospray ionization-mass spectrometry (ESI-MS)	Elucidation of biological mass, amino acid sequences, and modified structure of peptides and proteins	Ionized biological sample gives all about the mass, sequence and three-dimensional structure	Tsuchiya et al. (2020)
Systematic Evolution of Ligands by Exponential Enrichment (SELEX)	Isolate aptamers from different microbial targets, toxins, and chemical compound	Deciphering the protein-DNA sequence specificity to address the fundamental biological query	Liu et al. (2020)
Microchip Capillary Electrophoresis	Identification and separation of metabolites and their profiling	Highly applicable due to the requirement of less sample, short analysis time, high-throughput capability, low waste generation, and portability	Shankar et al. (2019)
2-D fluorescence difference gel electrophoresis (2D-DIGE)	Labeling and separation of one or more proteins by isoelectric focusing	Done with comparative proteomics study by separating complex protein into simple component	Vilasi et al. (2013)
Exponentially modified protein abundance index (emPAI)	Comparative proteomic analysis performed to estimate protein abundance	Easy to applied in multi-dimensional proteomic separation-MS/MS	Shinoda et al. (2010)
iTRAQ/LC-MS/MS	Identify total protein with their differential regulation	Widely used for quantitative proteomics, prediction of SM production	Martinez-Gomez et al. (2012)
Nano-flow liquid chromatography tandem mass spectrometry (nano-flow LC-MS/MS)	Proteolytically digested proteins isolated by 2-D gel electrophoresis	Robust tool in the qualitative and quantitative protein identification	Laatsch (2011)
Stable isotope labeling by amino acids in cell culture (Absolute SILAC)	Identify targeted labeled protein from a large set of samples	Applied for targeted protein quantification in various biomedical research and clinical practices	Wang et al. (2016)
LTQ-Orbitrap LC MS/MS	Identify and structurally characterize peptides in a highly complex sample mixture	More sensitive with higher accuracy then other proteomic tools	Narayanan et al. (2010)
Label-free LCMS/MS	Total protein profiling in a single run with their quantification	Fast, low-cost, rigorous, powerful tools for analyzing protein changes in large-scale proteomics studies	Jain et al. (2020)