Amino-terminal sequence and processing of the precursor of the leucine-specific binding protein, and evidence for conformational differences between the precursor and the mature form.

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Abstract
A 2.1-kilobase BglII DNA fragment from Escherichia coli containing livK, the gene coding for the leucine-specific binding protein, was cloned into the BamHI site of the plasmid vector pBR322. The DNA sequence of segments of the resulting plasmid, pOX7, established the location of the livK gene and the direction of its transcription. In vitro protein synthesis directed by pOX7 DNA yielded the MW 42,000 precursor of the leucine-specific binding protein and a small amount of the MW 39,000 mature protein. Continued incubation of the in vitro reaction mixture after DNase and RNase treatment resulted in additional processing. The DNA sequence of the beginning of livK suggested that 23 additional amino acid residues are present as an extension of the NH2 terminus of the mature protein. Amino acid sequence analysis established that the precursor has the predicted 23-residue extension. Proteolytic digestion studies with the precursor and mature forms of the leucine-specific binding protein indicate that there are conformational differences between the 2. This suggests a possible role for the signal sequence in determining the conformation of the binding protein precursor that is recognized by the membrane.