Cloning and sequencing of the pheP gene, which encodes the phenylalanine-specific transport system of Escherichia coli.
The phenylalanine-specific permease gene (pheP) of Escherichia coli has been cloned and sequenced. The gene was isolated on a 6-kb Sau3AI fragment from a chromosomal library, and its presence was verified by complementation of a mutant lacking the functional phenylalanine-specific permease. Subcloning from this fragment localized the pheP gene on a 2.7-kb HindIII-HindII fragment. The nucleotide sequence of this 2.7-kb region was determined. An open reading frame was identified which extends from a putative start point of translation (GTG at position 636) to a termination signal (TAA at position 2010). The assignment of the GTG as the initiation codon was verified by site-directed mutagenesis of the initiation codon and by introducing a chain termination mutation into the pheP-lacZ fusion construct. A single initiation site of transcription 30 bp upstream of the start point of translation was identified by the primer extension analysis. The pheP structural gene consists of 1,374 nucleotides specifying a protein of 458 amino acid residues. The PheP protein is very hydrophobic (71% nonpolar residues). A topological model predicted from the sequence analysis defines 12 transmembrane segments. This protein is highly homologous with the AroP (general aromatic transport) system of E. coli (59.6% identity) and to a lesser extent with the yeast permeases CAN1 (arginine), PUT4 (proline), and HIP1 (histidine) of Saccharomyces cerevisiae.
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J
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Department of Microbiology, University of Melbourne, Parkville, Victoria, Australia.
Wookey
P J
PJ
Pittard
A J
AJ
D000595:Amino Acid Sequence; D026905:Amino Acid Transport Systems; D026921:Amino Acid Transport Systems, Neutral; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics*; D001483:Base Sequence; D001692:Biological Transport; D002352:Carrier Proteins / Q000737:chemistry; D002876:Chromosomes, Bacterial; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D029968:Escherichia coli Proteins; D005798:Genes, Bacterial; D026901:Membrane Transport Proteins / Q000235:genetics*; D008969:Molecular Sequence Data; D009693:Nucleic Acid Hybridization; D016366:Open Reading Frames; D010649:Phenylalanine / Q000378:metabolism*; D014176:Protein Biosynthesis; D011487:Protein Conformation; D012329:RNA, Bacterial / Q000235:genetics; D015183:Restriction Mapping; D012689:Sequence Homology, Nucleic Acid; D014158:Transcription, Genetic
D026905:Amino Acid Transport Systems; D026921:Amino Acid Transport Systems, Neutral; C063201:AroP protein, E coli; D001426:Bacterial Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; D012329:RNA, Bacterial; C069320:phenylalanine-specific permease, E coli; D010649:Phenylalanine