Role of N-glycosylation in the expression of human band 3-mediated anion transport.
The human erythrocyte anion transporter (band 3; AE1) has a single N-linked glycosylation site at amino residue Asn-642. To investigate the functional role of the N-glycan in band 3 (b3) we have constructed mutant b3 cDNAs in which this residue has been replaced by Gly, Ser or Thr, and the expression of these mutants was examined in Xenopus oocytes. Chymotrypsin treatment of intact oocytes was used to assess surface b3. Similar amounts of cleavage were observed with both glycosylated and unglycosylated b3. Greater cleavage of b3 was obtained when human red cell glycophorin A (GPA) was co-expressed with either glycosylated or unglycosylated b3. The co-expression of GPA with either glycosylated or unglycosylated b3 increased the stilbene disulphonate-sensitive chloride transport into oocytes at low cRNA concentrations. In both the presence or absence of GPA, a higher b3-mediated chloride influx into oocytes was observed on expression of glycosylated b3 cRNA compared with similar amounts of unglycosylated b3 cRNA. We suggest that glycosylation is not essential for the expression of functional b3 in oocytes, but may play a role in enabling the protein to acquire its correct folding with the highest anion transport activity.
Department of Biochemistry, School of Medical Sciences, University of Bristol, UK.
J D
JD
Groves
M J
MJ
Tanner
D000818:Animals; D001457:Anion Exchange Protein 1, Erythrocyte / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001483:Base Sequence; D002918:Chymotrypsin / Q000494:pharmacology; D018076:DNA, Complementary / Q000235:genetics; D005260:Female; D006031:Glycosylation; D006596:Hexosaminidases / Q000494:pharmacology; D006801:Humans; D017136:Ion Transport; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000378:metabolism; D014176:Protein Biosynthesis; D014981:Xenopus
D001457:Anion Exchange Protein 1, Erythrocyte; D018076:DNA, Complementary; D006596:Hexosaminidases; D002918:Chymotrypsin