Journal of Biological Chemistry
Volume 277, Issue 45, 8 November 2002, Pages 42741-42747
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MEMBRANE TRANSPORT STRUCTURE FUNCTION AND BIOGENESIS
Two Highly Conserved Glutamate Residues Critical for Type III Sodium-dependent Phosphate Transport Revealed by Uncoupling Transport Function from Retroviral Receptor Function*

https://doi.org/10.1074/jbc.M207096200Get rights and content
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Type III sodium-dependent phosphate (NaPi) cotransporters, Pit1 and Pit2, have been assigned housekeeping Pi transport functions and suggested involved in chondroblastic and osteoblastic mineralization and ectopic calcification. Both proteins exhibit dual function, thus, besides being transporters, they also serve as receptors for several gammaretroviruses. We here show that it is possible to uncouple transport and receptor functions of a type III NaPicotransporter and thus exploit the retroviral receptor function as a control for proper processing and folding of mutant proteins. Thus exchanging two putative transmembranic glutamate residues in human Pit2, Glu55 and Glu575, with glutamine or with lysine severely impaired or knocked out, respectively, Pitransport function, but left viral receptor function undisturbed. Both glutamates are conserved in type III NaPi cotransporters, in fungal NaPi cotransporters PHO-4 and Pho89, and in other known or putative phosphate permeases from a number of species and are the first residues shown to be critical for type III NaPicotransport. Their putative transmembranic positions together with the presented data are consistent with Glu55 and Glu575 being parts of a cation liganding site or playing roles in conformational changes associated with substrate transport. Finally, the results also show that Pit2 retroviral receptor function per se is not dependent on Pit2 Pitransport function.

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Published, JBC Papers in Press, August 29, 2002, DOI 10.1074/jbc.M207096200

*

This work was supported in part by the Danish Medical Research Council (Grant 9802349), the Novo Nordisk Foundation, the Karen Elise Jensen Foundation, the Danish Cancer Society (Grant DP00092), and the Lundbeck Foundation.The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

The on-line version of this article (available at www.jbc.org) contains the alignments of 2.TM, 3.TM, and 9.TM sequences from Pit1, Pit2, and related proteins from the indicated species.

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Supported by a Scholarship from the Danish Cancer Society (Grant 190015).