Time for Multiple Extraction Methods in Proteomics? A Comparison of Three Protein Extraction Methods in the Eustigmatophyte Alga Microchloropsis gaditana CCMP526

Abstract

Proteomics is a crucial postgenomic biotechnology for functional and systems scale analyses in cell and integrative biology, not to mention clinical and precision medicine research. However, a fundamental requirement for an accurate examination of the protein complement of cells is an efficient method for extracting the proteins. This study reports on the evaluation of three protein extraction methods: trichloroacetic acid (TCA)-acetone, phenol, and TRIzol, in the eustigmatophyte alga Microchloropsis gaditana CCMP526 for proteomic analysis. M. gaditana is a potential candidate for algal-based biofuels. This industrially important strain is also rich in dietary oil and pigments and is used as feed in the aquaculture industry. Liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomic analysis was performed for proteins obtained using the three extraction methods and their effects were examined by the abundance ratio. Protein yield was higher using the TCA-acetone and phenol methods than with the TRIzol method. The TCA method was superior than other methods examined here in terms of protein coverage and abundance. Subcellular localization of the identified proteins revealed no significant difference among the extraction methods. Importantly, each method revealed a unique set of proteins. To the best of our knowledge, this is the first report on evaluation of protein extraction methods for the proteomic analysis of M. gaditana CCMP526. These observations underscore the importance of using multiple protein extraction methods for comprehensive proteome coverage, as the field of proteomics edges toward diverse applications in biofuels, aquaculture industry, marine biology, and agriculture.

Keywords: Microchloropsis; mass spectrometry; postgenomic biotechnology; protein extraction; proteomics.