Identification of mitochondrial proteins on two-dimensional electrophoresis gels of extracts of adult Drosophila melanogaster

Abstract

Abstract Several hundred proteins have been resolved on two-dimensional gels of extracts of [35S]methionine-labeled adult Drosophila melanogaster. 27 of these polypeptides disappear from the gel pattern after feeding the K+ ionophore nonactin. These proteins have been identified as mitochondrial, since the two-dimensional gel pattern of extracts of isolated mitochondria correlates well with the pattern of the proteins missing from that of nonactin-treated flies. Nine new proteins also appear on the two-dimensional gels of the extracts from the nonactin-treated flies. Apparently, these nine proteins are precursors of the mature mitochondrial forms. These particular data support the concept that processing of many of the cytoplasmically synthesized mitochondrial proteins requires a specific membrane potential, and that some of these proteins are modified intramitochondrially. However, using [35S]methionine incorporation techniques, not all labeled polypeptides disappear from mitochondria during such treatment. Feeding similarly radiolabeled flies with chloramphenicol, an inhibitor of mitochondrial protein synthesis, results in the disappearance of only one protein from the gel pattern with the concurrent appearance of a ‘new’ high-molecular-weight polypeptide. Collectively, these data show that a specific group of [35S]methionine-labeled mitochondrial proteins can be identified by selective inhibition of mitochondrial function in whole cell protein maps of adult D. melanogaster.