Biochemical characterization of a mutant lipoprotein of Escherichia coli

Abstract

A lipoprotein mutant of E. coli K-12 has been characterized. The mutant lipoprotein was found to differ from the wild-type lipoprotein in the following respects: (i) it is present in an appreciable amount in the soluble fraction (275,000 X g supernatant); (ii) it lacks the covalently-linked diglyceride; (iii) it contains an unmodified cysteine which can be carboxymethylated in vitro; (iv) it undergoes dimerization and the dimer can be converted into monomeric form by reduction with 2-mercaptoethanol; (v) both the monomeric form and especially the dimeric form of the mutant lipoprotein migrate more slowly than the corresponding forms of wild-type lipoprotein in sodium dodecyl sulfate/urea polyacrylamide gel electrophoresis; and (vi) the mutant lipoprotein is not assembled into the murein sacculi, and this results in a greatly reduced amount of bound-form lipoprotein in the mutant. These data strongly suggest that the mutation has affected the primary structure of lipoprotein, in such a way that it is not modified normally, leading to the production of a structurally-altered lipoprotein deficient in covalently-linked lipid as well as a defective assembly of the altered lipoprotein into the rigid layer of the cell envelope.