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Journal of Biochemistry Advance Access originally published online on January 3, 2009
Journal of Biochemistry 2009 145(4):437-443; doi:10.1093/jb/mvn181
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© The Authors 2009. Published by Oxford University Press on behalf of the Japanese Biochemical Society. All rights reserved

Identification and Functional Characterization of Rat Riboflavin Transporter 2

Syunsuke Yamamoto*, Katsuhisa Inoue*, Kin-ya Ohta, Rui Fukatsu, Jun-ya Maeda, Yukihiro Yoshida and Hiroaki Yuasa{dagger}

Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Nagoya City University, Nagoya 467-8603, Japan

{dagger}To whom correspondence should be addressed. Tel: +81-52-836-3423, Fax: +81-52-836-3423, E-mail: yuasa{at}phar.nagoya-cu.ac.jp

Received October 22, 2008; Accepted December 15, 2008


  Abstract

We have newly identified rat riboflavin transporter 2 (rRFT2) and its human orthologue (hRFT2), and carried out detailed functional characterization of rRFT2. The mRNA of rRFT2 was highly expressed in jejunum and ileum. When transiently expressed in human embryonic kidney (HEK) 293 cells, rRFT2 could transport riboflavin efficiently. Riboflavin transport mediated by rRFT2 was Na+-independent but moderately pH-sensitive, being more efficient in acidic conditions than in neutral and basic conditions. Kinetic analysis indicated that rRFT2-mediated riboflavin transport was saturable with a Michaelis constant (Km) of 0.21 µM. Furthermore, it was specifically and strongly inhibited by lumiflavin, flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD), and to a lesser extent by amiloride. Such ability to transport riboflavin in a specific manner, together with its high expression in the small intestine, indicates that RFT2 may play a role in the intestinal absorption of riboflavin.

Key Words: absorption, intestine, riboflavin, RFT2, transporter

Abbreviations: BSP, sulfobromophthalein; DIDS, 4,4'-diisothiocyanostilbene-2,2'-disulfonate; EDTA, ethylenediaminetetraacetic acid; FAD, flavin adenine dinucleotide; FMN, flavin mononucleotide; GAPDH, glyceraldehyde-3-phosphate dehydrogenase; GFP, green fluorescent protein; HEK, human embryonic kidney; HEPES, 4-(2-hydroxyethyl)piperazine-1-ethanesulfonic acid; MES, 2-(N-morpholino)ethanesulfonic acid; PCR, polymerase chain reaction; RFT, riboflavin transporter; RT, reverse transcription; SDS, sodium dodecyl sulphate; TRIS, tris(hydroxymethyl)aminomethane

*These authors contributed equally to this work.


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