Title : The human colonic thiamine pyrophosphate transporter (
hTPPT ) is a
glycoprotein and N-linked glycosylation is important for its function
Abstract :
- The recently identified human thiamine pyrophosphate transporter ( hTPPT ; product of the SLC44A4 gene) is responsible for absorption of the microbiota-generated TPP in the large intestine
- The hTPPT is highly expressed in the colon, but not in other regions of the intestinal tract and is localized exclusively at the apical membrane domain of epithelia
- The hTPPT protein is predicted to have multiple TM domains with a number of putative N-glycosylation sites , but it is not known if the protein is actually glycosylated, and if so at which site, and their role in the functionality of the transporter
- Using several approaches including inhibiting de novo N-glycosylation in human colonic epithelial NCM460 cells with tunicamycin as well as enzymatic de-glycosylation, we show that the hTPPT protein is, indeed, a glycoprotein
- Glycosylation of hTPPT was shown, by mean of site-directed mutagenesis, to occur at Asn(69), Asn(155), Asn(197), Asn(393), and Asn(416)
- However, only N-glycosylation at Asn(69), Asn(155), and Asn(393) appeared to be important for transporter functionality possibly through an effect on protein conformation and/or interaction with its ligand (but not through changes in expression at the cell membrane as determined by live cell confocal imaging)
- Results of this study showed, for the first time, that the hTPPT is glycosylated and that N-linked glycosylation occurs at multiple sites with some of them being important for function
- The results also provide an indirect support for a membrane topology for hTPPT with 10 transmembrane domains as predicted by the TMHMM transmembrane helixes prediction program
Output (sent_index, trigger,
protein,
sugar,
site):
- 0. glycoprotein, , glycoprotein, -, -
- 3. N-glycosylation, , -, -, sites
- 3. glycosylated, , protein, -, -
- 4. glycoprotein, , glycoprotein, -, -
- 4. glycoprotein, , hTPPT protein, -, -
- 5. Glycosylation, , hTPPT, -, -
- 6. N-glycosylation, , -, -, Asn(69), Asn(155), and Asn(393)
- 7. glycosylated, , hTPPT, -, -
- 7. glycosylation, , -, -, sites
Output(Part-Of) (sent_index,
protein,
site):
- 3. hTPPT protein, domains
*Output_Site_Fusion* (sent_index,
protein,
sugar,
site):
- 6. hTPPT, -, Asn(69), Asn(155), and Asn(393)