CLONING AND ANALYSIS OF THE SHIA GENE, WHICH ENCODES THE SHIKIMATE TRANSPORT-SYSTEM OF ESCHERICHIA-COLI K-12

In Escherichia coli K-12, the shiA gene is involved in the uptake of s hikimate. This gene has been cloned and its nucleotide sequence determ ined. The gene is predicted to encode a protein of 438 amino acids and lies adjacent to the amn gene. The hydropathy profile and the amino a cid sequence indicate that the ShiA protein is a polytopic membrane pr otein that shows a homology with members of the major facilitator supe rfamily of transport proteins. Recombining an inactive form of the clo ned gene into the chromosome creates mutants unable to transport shiki mate. Introducing a wild-type gene on a multicopy plasmid into a shiA mutant restores the ability to transport shikimate. When this multicop y shiA plasmid is introduced into an aroE strain, this strain is now a ble to grow with shikimate as the aromatic supplement, consistent with the notion that dehydroshikimate (DHS) accumulated in an aroE strain prevents uptake of shikimate by competition. Expression of the shiA ge ne does not appear to be regulated by the TyrR protein, a repressor/ac tivator that controls the expression of other genes involved with the biosynthesis or transport of the aromatic amino acids. (C) 1998 Elsevi er Science B.V.