The human proton-dependent dipeptide transporter (PEPT1, gene SLC15A1) is important for intestinal absorption of di-and tripeptides and a variety of peptidomimetic compounds. Using a DNA polymorphism discovery panel of 44 ethnically diverse individuals, nine nonsynonymous and four synonymous coding-region single-nucleotide polymorphisms (SNPs) were identified in PEPT1. HeLa cells were transiently transfected with plasmids constructed by site-directed mutagenesis for each of the nine nonsynonymous variants. Quantitative polymerase chain reaction showed that the mRNA transcription level of all of the mutants was comparable with the mRNA transcription level of the reference sequence in transfected HeLa cells. Functional analysis in transiently transfected HeLa cells revealed that all nonsynonymous variants retained similar pH-dependent activity and K t values for [glycyl-1,2-14 C]glycylsarcosine (GlySar) uptake as the reference PEPT1. In addition, a group of seven peptide-like drugs showed inhibitory effect on Gly-Sar uptake by these variants comparable with the reference, suggesting conserved drug recognition. Of the nine nonsynonymous SNPs, a single SNP (P586L) demonstrated significantly reduced transport capacity as evidenced by a much lower V max value. This was consistent with lower immunoactive protein level (Western analysis) and lower plasma membrane expression (immunocytochemical analysis). Therefore, Pro 586 may have profound effect on PEPT1 translation, degradation, and/or membrane insertion.

Human proton-dependent peptide transporters, PEPT1 and PEPT2, mediate the cellular uptake of di- and tripeptides as well as a variety of drug molecules. Although PEPT1 and PEPT2 have been cloned from many species, there are no data available for monkey, an important pharmacological and preclinical species in drug development. In this study, it was first verified that monkey intestine transports a model dipeptide, Gly-Sar, in a proton-dependent manner (0.30 +/- 0.05 pmol cm(-2) s(-1) at pH 6.0 and 0.10 +/- 0.03 pmol cm(-2) s(-1) at pH 7.4) in the absorptive direction, presumably by monkey PEPT1. RT-PCR and rapid amplification of cDNA ends (RACE) were then used to clone monkey PEPT1 and PEPT2. Monkey PEPT1 (2127 bp and 708 amino acids) was found to be >94 and > 92% identical to human PEPT1 at the cDNA and amino acid level, respectively. Monkey PEPT2 (2190 bp and 729 amino acids) was found to be > 97% identical to human PEPT2 at both the cDNA and amino acid levels. Functional comparison of human and monkey peptide transporters expressed in HeLa cells suggested that functionalities of PEPT1 and PEPT2 were largely conserved in terms of Gly-Sar uptake kinetics and inhibitor specificity (for most tested substrates). Finally, Northern and RT-PCR analyses revealed some differences in tissue mRNA levels of peptide transporters between human and monkey.