Abstract
This protocol describes a rapid protein extraction method for the archaeon Candidatus Vulcanisaeta moutnovskia, which can be also implemented for other archaea. The utilization of two different methods for protein extraction constitute the main step of the protocol. Method I involves the extraction with a multi-chaotropic lysis buffer containing a non-denaturing zwitterionic detergent, most efficient for extracting cytosolic proteins. Method II involves a denaturing anionic detergent allowing total disruption of the membranes and capable of extracting both membrane (hydrophobic) and non-membrane (water-soluble, hydrophilic) proteins. The big advantage of the methods is to use general laboratory chemicals to make powerful extraction buffers, resulting in high quality and quantity of proteins. The methods probably are usable for any other archaea or microbial cells, and takes about 14-22 h. Following extraction and further protein digestion, 1D-nano Liquid Chromatography Electrospray Ionization Tandem Mass Spectrometric (LC ESI-MSMS) analysis with Triple TOF 5600 and Orbitrap technologies were used for protein identification and further quantification.