Characterisation of the H(+)/peptide cotransporter of eel intestinal brush-border membranes.
H(+)/peptide cotransport in brush-border membrane vesicles (BBMVs) from eel (Anguilla anguilla) intestine was studied by measuring d-[(3)H]-phenylalanyl-l-alanine uptake and by monitoring peptide-dependent intravesicular acidification using the pH-sensitive dye Acridine Orange. d-[(3)H]-phenylalanyl-l-alanine influx was greatly stimulated by an inside-negative membrane potential and enhanced by an inwardly directed H(+) gradient. In parallel, vesicular H(+) influx was significantly increased in the presence of extravesicular d-phenylalanyl-l-alanine or a series of glycyl and l-prolyl peptides. H(+)/peptide cotransport displayed saturable kinetics involving a single carrier system with apparent substrate affinities of 0.9-2.6 mmol l(-1) depending on the particular peptide. All substrates tested competed with this system. Pre-incubation of BBMVs with dipeptides prevented diethylpyrocarbonate inhibition of transport activity, suggesting that the substrates mask histidine residues involved in the catalytic function of the transporter. Using human PepT1-specific primers, a reverse transcription-polymerase chain reaction (RT-PCR) signal was detected in eel intestine. Our results suggest that, in eel intestine, a brush-border membrane 'low-affinity'-type H(+)/peptide cotransport system is present that shares kinetic features with the mammalian intestinal PepT1-type transporters.
Verri
T
T
Laboratory of General Physiology, Department of Biology, University of Lecce, Strada Provinciale Lecce-Monteroni, I-73100 Lecce, Italy. physiol@ultra5.unile.it
Maffia
M
M
Danieli
A
A
Herget
M
M
Wenzel
U
U
Daniel
H
H
Storelli
C
C
D000811:Anguilla / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D001693:Biological Transport, Active; D015820:Cadherins; D002352:Carrier Proteins / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers / Q000235:genetics; D004151:Dipeptides / Q000378:metabolism; D006801:Humans; D007413:Intestinal Mucosa / Q000378:metabolism*; D007700:Kinetics; D026901:Membrane Transport Proteins; D008871:Microvilli / Q000378:metabolism; D000074011:Peptide Transporter 1; D010455:Peptides / Q000378:metabolism; D011522:Protons; D027981:Symporters
C490584:CDH17 protein, human; D015820:Cadherins; D002352:Carrier Proteins; D017931:DNA Primers; D004151:Dipeptides; D026901:Membrane Transport Proteins; D000074011:Peptide Transporter 1; D010455:Peptides; D011522:Protons; C082186:SLC15A1 protein, human; D027981:Symporters; C067802:intestinal peptide-proton cotransporter; C039552:phenylalanylalanine