title: Direct evidence for efficient transport and minimal metabolism of L-cephalexin by oligopeptide transporter 1 in budded baculovirus fraction.
abstract: The oligopeptide transporter PEPT1 (SLC15A1) is responsible for absorption of peptidic nutrients in the small intestine. Although the L-diastereomer of the beta-lactam antibiotic cephalexin (L-cephalexin) is likely to be transported by PEPT1, there has been no direct demonstration of PEPT1-mediated L-cephalexin transport. Indeed, after the incubation with L-cephalexin, the intact form of L-cephalexin has not been identified inside vesicles/proteoliposomes prepared from brush border membrane of intestinal epithelial cells or cultured cell lines exogenously transfected with PEPT1 gene. Thus, it appears that L-cephalexin is rapidly metabolized by PEPT1 or PEPT1-associated proteins. Here, we attempted to verify whether L-cephalexin is transported by PEPT1 and whether it is hydrolyzed by PEPT1 itself, by using budded baculovirus expressing PEPT1 protein. Marked uptake of L-cephalexin in PEPT1-expressing budded baculovirus, compared with wild-type virus, indicated that L-cephalexin is a substrate for PEPT1. The uptake was found to be pH sensitive, and was strongly inhibited by the D-diastereomer of cephalexin and glycylsarcosine, but not by glycine. Thus, L-cephalexin is transported by PEPT1 itself. Upon the transport of both L- and D-cephalexin by PEPT1, dose-dependent membrane depolarization was observed; the EC(50) values of 0.18 and 2.9 mM, respectively, indicate that the affinity of L-cephalexin for PEPT1-mediated transport is much higher than that of the D-diastereomer. On the other hand, the L-cephalexin metabolite 7-aminodesacetoxycephalosporanic acid was not detected in PEPT1-expressing or wild-type virus at either pH 6.0 or 7.4. We conclude that L-cephalexin is transported by PEPT1 with high affinity, but is not metabolized by PEPT1 itself.
authors: Faculty of Pharmacy, Institute of Medical, Pharmaceutical and Health Sciences, Kanazawa University, Kanazawa, Japan.
authors: Keisuke
authors: K
authors: Mitsuoka
authors: Ikumi
authors: I
authors: Tamai
authors: Yasushi
authors: Y
authors: Morohashi
authors: Yoshiyuki
authors: Y
authors: Kubo
authors: Ryoichi
authors: R
authors: Saitoh
authors: Akira
authors: A
authors: Tsuji
authors: Yukio
authors: Y
authors: Kato
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000737:chemistry / Q000378:metabolism*; D016367:Baculoviridae / Q000378:metabolism*; D001692:Biological Transport; D002460:Cell Line; D002506:Cephalexin / Q000737:chemistry / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007413:Intestinal Mucosa / Q000378:metabolism; D008564:Membrane Potentials / Q000187:drug effects; D000074011:Peptide Transporter 1; D013237:Stereoisomerism; D013379:Substrate Specificity; D027981:Symporters / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism*
chemical_list: D000900:Anti-Bacterial Agents; D000074011:Peptide Transporter 1; C082186:SLC15A1 protein, human; D027981:Symporters; D002506:Cephalexin