Quantitative Proteomics Analysis Using MS~(E) Data Acquisition of Chlamydomonas reinhardtii Under Copper-Deficiency Conditions

摘要

Copper is a micronutrient necessary for several important biochemical processes including respiration, photosynthesis and oxidative stress management. Chlamydomonas is one reference system for studying the metabolic consequences of copper-deficiency. Quantitative proteomic approaches may inform us about proteins whose abundance is impacted by copper-deficiency. Label-free, MS~(E) based quantification has been performed to assess changes to the proteome. MS~(E) is a data acquisition mode that facilitates the simultaneous collection of qualitative and quantitative information, in a data independent fashion and without the use of isotope labeling. The instrument continuously alternates between a low-energy scanning mode (MS) for accurate mass peptide precursor identification, and an elevated-energy mode (MS~(E); by raising the energy in the collision cell of the QToF) for generation of accurate mass multiplex peptide fragmentation data. From the alternate scanning acquisition method, products produced from any given precursor will have the same chromatographic profile and retention time as the originating precursor ion. Absolute quantification is based on a relationship between MS signal response and protein concentration, and requires an internal digested standard of known quantity.