Cloning of the tyrP gene and further characterization of the tyrosine-specific transport system in Escherichia coli K-12.
The tyrP gene which codes for a component of the tyrosine-specific transport system of Escherichia coli has been cloned on a 2.8-kilobase insert into plasmid pBR322. Transposon mutagenesis, using Tn1000, indicates that the tyrP+ gene is at least 1.1 kilobase in length. Labeling of the tyrP protein in maxicells with [35S]methionine indicates an apparent molecular weight of ca. 24,500. Sedimentation analysis reveals that the tyrP protein is associated with the cell membrane and is not free in the cytoplasm or periplasm. Strains with many copies of the tyrP+ gene show an enhanced uptake of tyrosine, but the expression of the system is still modulated by tyrosine and phenylalanine in the presence of the tyrR+ regulator protein. Accumulated radioactive tyrosine is rapidly effluxed by the addition either of energy uncouplers or of excess nonradioactive tyrosine, indicating that the transport system is energized by the proton motive force and that the internal pool is readily exchangeable. The effect of increasing expression of the tyrP gene on the steady-state level of tyrosine accumulated by cells indicates that although the transport system may be dependent on the proton motive force to drive uptake, the system never reaches thermodynamic equilibrium with it.
D001483:Base Sequence; D001693:Biological Transport, Active; D003001:Cloning, Molecular; D004262:DNA Restriction Enzymes; D004251:DNA Transposable Elements; D004926:Escherichia coli / Q000235:genetics*; D005796:Genes; D005798:Genes, Bacterial; D005838:Genotype; D007700:Kinetics; D008970:Molecular Weight; D009154:Mutation; D010641:Phenotype; D010957:Plasmids; D013462:Sulfur Radioisotopes; D014443:Tyrosine / Q000378:metabolism*
D004251:DNA Transposable Elements; D013462:Sulfur Radioisotopes; D014443:Tyrosine; D004262:DNA Restriction Enzymes