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Please use this identifier to cite or link to this item: http://hdl.handle.net/10277/684

Authors: Renna, Maria Daniela
Internal Tutor: PERES, ANTONIO
Title: Structure-function relationships in the oligopeptide transporter PepT1.
Abstract: The proton-dependent di- and tripeptide transporter PepT1 represents the major route of dietary amino acid intake in the intestine of many species. This transporter belongs to the solute carrier family SLC15 and because of its electrogenic properties it may be studied both through electrophysiological and radiotracer uptake experiments. In this work some functional and structural aspects of PepT1 have been investigated at the molecular level. In addition to the physiological relevance, understanding of the details of its mechanisms of operation is important since PepT1 appears to be involved in the absorption of many important, orally administered, drugs such as antibiotics, angiotensin-converting inhibitors, and antiviral agents. The electrophisyiological and biophysical properties of PepT1 expressed in Xenopus oocytes were investigated with two-electrode voltage-clamp. Most of the above functional observations were derived from uptake data, in absence of control of the membrane voltage, or from electrophysiological measurement of steady transport currents, in the presence of a dipeptide substrate. Important additional informations regarding the transport mechanism may arise from measurement of presteady-state currents, the electrophysiological signals that can be observed in absence of organic substrate, and that represent the first steps in the transport cycle. A unified kinetic model for PepT1 has been devised that can describe the different characteristics of the isoforms of different species, with respect to both presteady-state and transport-associated currents. Mutational studies have provided significant evidence for the functional role of some residues in PepT1. Particularly Arg282 and Asp341, in the transmembrane domain 7 and 8 of the transporter, have been reported to form a charge pair that may break and reform during the transport cycle. The attention has been focused on these two oppositely charged aminoacids to better understand their functional role in the absorption pathway. Finally, the functional and structural basis of reverse operation of wild-type and mutated forms of PepT1 have been studied. Mutants in the putative charge pair residues Arg282 and Asp341 of rabbit PepT1 have been shown exhibit properties useful to better understand the possibility of reverse transport. This reversed mode of operation may be either the effect or the cause of abnormal or pathological conditions.
Keywords: PepT1, transporter, oligopeptides, proton-dependence, modeling, electrostatic gates, reverse operation, electrophysiology.
Issue Date: 2011
Language: eng
Doctoral course: Biologia Cellulare e Molecolare
Academic cycle: 24
Publisher: Università degli Studi dell'Insubria
Citation: Renna, M.D.Structure-function relationships in the oligopeptide transporter PepT1. (Doctoral Thesis, Università degli Studi dell'Insubria, 2011).

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