COMPOUND MISSENSE MUTATIONS IN THE SODIUM D-GLUCOSE COTRANSPORTER RESULT IN TRAFFICKING DEFECTS/

Background & Aims: Defects in the Na+-dependent glucose transporter (S GLT1) are associated with the disorder glucose-galactose malabsorption , characterized by severe diarrhea. This study focused on a unique pro band with glucose-galactose malabsorption who was investigated 30 year s ago, and the aims of the study were to identify mutations in the SGL T1 gene and to determine the defect in sugar transport. Methods: Mutat ions were identified by sequencing, and each mutant protein was then s tudied using a Xenopus oocyte heterologous expression system. Analysis included Western, freeze fracture, radiotracer uptake, and electrophy siological assays. Results: Two heterozygous missense mutations (Cys35 5Ser and Leu147Arg) were identified that entirely eliminated Na+/sugar cotransport activity. Western blot analysis showed that the levels of both mutant proteins in the oocyte were comparable to wild-type SGLT1 , but no complex glycosylation was detected. No SGLT1 charge movements were observed with the mutant proteins, and freeze fracture data show ed that neither mutant protein reached the plasma membrane. Conclusion s: The Cys355Ser and Leu147Arg mutations eliminate the Na+/sugar cotra nsport by blocking the transfer of SGLT1 protein from the endoplasmic reticulum to the plasma membrane. This is consistent with earlier stud ies on phlorizin binding to the brush border membrane of duodenal biop sy specimens from this patient.