Cloning of the genes for polyamine transport proteins and regulation of its expression

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Polyamine uptake is known to increase during cell proliferation and to be energy dependent. We recently succeeded in obtaining three clones of polyamine transport genes (pPT104, pPT79, and pPT71) in E. coli. Spermidine uptake was catalyzed by the system encoded by pPT104, and putrescine uptake was catalyzed by the systems encoded by pPT104 and pPT79 in the absence of spermidine. Putrescine uptake by the pPT79 system was not influenced by spermidine, whereas that by the pPT104 system was greatly inhibited. These two systems consisted of four kinds of proteins: a periplasmic substrate binding protein (potD and potF proteins for pPT104 and pPT79, respectively), a membrane associated protein having the nucleotide binding site (potA and potG proteins) and two other membrane proteins having 6 putative transmembrane spanning segments (potB and C proteins for pPT104, and potH and I proteins for pPT79). The polymine uptake was shown to be both ATP- and periplasmic substrate binding protein-dependent, using membrane vesicles and E. coli mutants lacking Fo, F_0, F_1-ATPase. Excretion of putrescine was catalyzed by pPT71. The clone encoded Mr 46K protein which consisted of 12 putative transmembrane spanning segments linked by hydrophilic segments of variable length. The protein was an antiport protein between putrescine and ornithine (or lysine). The transport activity was not disturbed by the inhibitors of energy production such as KCN and carbonylcyanide m-chlorophenylhydrazone (CCCP).