Submitted on December 14, 2007
Accepted on April 3, 2008
Functional characterization, localization and molecular identification of rabbit intestinal N-amino acid transporter
Jamilur R. Talukder1, Ramesh Kekuda2, Prosenjit Saha2, Puttur D Prasad3, Vadivel Ganapathy4, and Uma Sundaram2*
1 Medicine, West Virginia University School of Medicine, Morgantown, West Virginia, United States
2 Department of Medicine, West Virginia University Medical Center, Morgantown, West Virginia, United States
3 Med Coll Georgia, United States
4 Department of Biochemistry & Molecular Biology, Medical College of Georgia, Augusta,, Georgia, United States
* To whom correspondence should be addressed. E-mail: usundaram{at}hsc.wvu.edu.
We have characterized the Na-glutamine co-transporter in the rabbit intestinal crypt cell BBMV. Using substrate specificity experiments it was shown that crypt cell glutamine uptake is mediated by system N. Real-time PCR experiments showed that SN2 (SLC38A5) mRNA is more abundant in crypt cells as compared to SN1 (SLC38A3) indicating that SN2 is the major glutamine transporter present in the apical membrane of the crypt cells. SN2 cDNA was obtained by screening a rabbit intestinal cDNA library using human SN1 as probe. Rabbit SN2 cDNA encompassed a 473 amino acid long open-reading frame. SN2 protein displayed 87% identity and 91% similarity to human SN2. Functional characterization studies of rabbit SN2 were performed using vaccinia virus mediated transient expression system. Substrate specificity of the cloned transporter was identical to that of SN2 described in the literature and matched well with substrate specificity experiments performed using crypt cell BBMV. Cloned rabbit SN2, analogous to its human counterpart, is Li+ tolerant. Hill coefficient for Li+ activation of rabbit SN2-mediated uptake was 1. Taken together, functional data from the crypt cell BBMV and the cloned SN2 cDNA indicate that the crypt cell glutamine transport is most likely mediated by SN2.
