Structural requirements for the stability and microsomal transport activity of the human glucose 6-phosphate transporter

Deficiencies in glucose 6-phosphate (G6P) transporter (G6PT), a 10-helical endoplasmic reticulum transmembrane protein of 429 amino acids, cause glyco gen storage disease type Ib. To date, only three missense mutations in G6PT have been shown to abolish microsomal G6P transport activity. Here, we rep ort the results of structure-function studies on human G6PT and demonstrate that 15 missense mutations and a codon deletion (Delta F93) mutation aboli sh microsomal G6P uptake activity and that two splicing mutations cause exo n skipping. While most missense mutants support the synthesis of G6PT prote in similar to that of the wild-type transporter, immunoblot analysis shows that G20D, Delta F93, and I278N mutations, located in helix 1, 2, and 6, re spectively, destabilize the G6PT. Further, we demonstrate that G6PT mutants lacking an intact helix 10 are misfolded and undergo degradation within ce lls. Moreover, amino acids 415-417 in the cytoplasmic tail of the carboxyl- domain, extending from helix 10, also play a critical role in the correct f olding of the transporter. However, the last 12 amino acids of the cytoplas mic tail play no essential role(s) in functional integrity of the G6PT. Our results, for the first time, elucidate the structural requirements for the stability and transport activity of the G6PT protein.