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Two‐dimensional electrophoresis of the fatty acid binding protein from human heart: Evidence for a thiol group which can form an intermolecular disulfide bond
Author(s) -
Nielsen Søren U.,
Vorum Henrik,
Spener Friedrich,
Brodersen Rolf
Publication year - 1990
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.1150111017
Subject(s) - isoelectric point , fatty acid binding protein , isoelectric focusing , chemistry , dimer , biochemistry , gel electrophoresis , polyacrylamide gel electrophoresis , gene isoform , thiol , electrophoresis , enzyme , gene , organic chemistry
A 100 000 g supernatant from human heart muscle, containing cytosolic proteins with some contaminating plasma proteins, was analyzed for fatty acid binding protein (FABP) by two‐dimensional electrophoresis (2‐DE) using isoelectric focusing under nondenaturing conditions in the first dimension. FABP purified from human heart muscle was found to comigrate with a major spot in 2‐DE gels of the supernatant. This spot was comparable with those of the myoglobins in staining intensity. When purified FABP was charged with [H]palmitate and subjected to nondenaturing 2‐DE, radioactivity always comigrated with this protein. Under denaturing and reducing conditions in the second dimension, FABP was found to have a p I of 5.3 and an apparent molecular weight of 15 000. Isoforms of FABP, reported here for the first time to occur in human heart muscle, were observed as minor spots focusing at pH 5.1 and 5.7. When electrophoresis in the second dimension was carried out under denaturing but nonreducing conditions, an additional protein appeared at pH 5.3 with an apparent molecular weight of about 30 000. This protein was identified as a dimer of FABP and evidence for the involvement of an intermolecular disulfide bond in this dimerization is presented.