Two-dimensional gel database of human breast carcinoma cell expressed proteins: An update

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Abstract
Previously, we reported a two‐dimensional gel map and database with molecular weight/isoelectric point (Mr/pI) loci for 22 proteins expressed in the breast carcinoma cell line, MDA‐MB231 (Rasmussen et al., Electrophoresis 1997, 18, 588–598). Here we update this database with Mr/pI loci for a further nine cytoplasmic proteins and three Triton X‐114 solubilised membrane proteins from MDA‐MB231 cells. In addition, a novel protein, previously represented only in expressed sequence tag (EST) databases, has been identified as a Triton X‐114 soluble protein and assigned an Mr/pI locus. During the course of isolating proteins from the Triton X‐114 fraction, we compared recoveries of proteins in sodium dodecyl sulphate‐polyacrylamide gel electrophoresis (SDS‐PAGE) gels after isoelectric focusing (IEF) using either immobilised pH gradients or carrier ampholytes. In these experiments, a significantly higher proportion of membrane proteins were visible in SDS‐polyacrylamide gels after the use of carrier ampholytes for the first dimension. We also report our mass spectrometric‐based procedure for identifying two‐dimensional electrophoresis (2‐DE) gel‐resolved proteins, combining in‐gel enzymatic digestion, 0.2 mm internal diameter (ID) capillary column reversed‐phase high‐performance liquid chromatography (RP‐HPLC) peptide mapping and electrospray ionisation — ion trap — mass spectrometry.