Proteomic basis for the possible use of lymphocytes for metabolic screenings

Summary.

The advent of proteomics has provided a tool for the concomitant identification and determination of a large series of proteins using two-dimensional gel electrophoresis with subsequent mass spectrometrical analysis. We tried an approach to analyse the high abundance enzyme proteome of a lymphocytic cell line.

Immortalised lymphocytes were grown in RPMI 1640 in the presence of glutamine, harvested and the 100,000 × g supernatant of the homogenate was applied on two-dimensional gel electrophoresis with subsequent in-gel digestion of protein spots and MALDI-TOF (Matrix-associated laser desorption/ionization mass spectroscopy) analysis of resulting peptides using specific software.

A series of 57 metabolic enzymes were identified including enzymes of carbohydrate, amino acid, purine and intermediary metabolism.

We are presenting a tool for the analysis of metabolic systems including enzyme deficiencies at the protein level with the advantage of unambiguous identification of proteins and thus complementing enzyme activity determinations.