Quantitative Mass Spectrometry by SILAC in Haloferax volcanii.

Abstract

The development of mass spectrometry (MS)-based proteomics methods has been critical in providing new insight about cellular processes and adaptations in all domains of life. While traditional MS-based methods are not inherently quantitative, technologies are now available to overcome this limitation. Of note, stable isotope labeling of amino acids in cell culture (SILAC) is reported as a reliable tool to label proteomes for quantitative MS-based proteomics that is accurate and flexible for multiplexing. The isotopically labeled lysine and arginine are easily incorporated into the proteome of cells auxotrophic for these amino acids. Microorganisms of the domain Archaea provide a fascinating alternative to understanding cellular adaptations and responses to environmental stresses. However, the availability of preferred SILAC-based quantitative analyses is limited. This protocol describes the use of SILAC to quantitatively analyze the proteome of Haloferax volcanii, a mesophilic halophilic archaeon that is easy to grow and requires no special equipment to maintain.