title: Glycosylation increases the thermostability of human aquaporin 10 protein.
abstract: Human aquaporin10 (hAQP10) is a transmembrane facilitator of both water and glycerol transport in the small intestine. This aquaglyceroporin is located in the apical membrane of enterocytes and is believed to contribute to the passage of water and glycerol through these intestinal absorptive cells. Here we overproduced hAQP10 in the yeast Pichia pastoris and observed that the protein is glycosylated at Asn-133 in the extracellular loop C. This finding confirms one of three predicted glycosylation sites for hAQP10, and its glycosylation is unique for the human aquaporins overproduced in this host. Nonglycosylated protein was isolated using both glycan affinity chromatography and through mutating asparagine 133 to a glutamine. All three forms of hAQP10 where found to facilitate the transport of water, glycerol, erythritol, and xylitol, and glycosylation had little effect on functionality. In contrast, glycosylated hAQP10 showed increased thermostability of 3-6 °C compared with the nonglycosylated protein, suggesting a stabilizing effect of the N-linked glycan. Because only one third of hAQP10 was glycosylated yet the thermostability titration was mono-modal, we suggest that the presence of at least one glycosylated protein within each tetramer is sufficient to convey an enhanced structural stability to the remaining hAQP10 protomers of the tetramer.
authors: Department of Chemistry/Biochemistry, University of Gothenburg, P. O. Box 462, SE-405 30 Göteborg, Sweden.
authors: Fredrik
authors: F
authors: Öberg
authors: Jennie
authors: J
authors: Sjöhamn
authors: Gerhard
authors: G
authors: Fischer
authors: Andreas
authors: A
authors: Moberg
authors: Anders
authors: A
authors: Pedersen
authors: Richard
authors: R
authors: Neutze
authors: Kristina
authors: K
authors: Hedfalk
mesh_terms: D020346:Aquaporins / Q000737:chemistry* / Q000378:metabolism; D001665:Binding Sites; D001692:Biological Transport; D006031:Glycosylation; D006358:Hot Temperature; D006801:Humans; D010843:Pichia / Q000235:genetics; D055550:Protein Stability
chemical_list: C438667:AQP10 protein, human; D020346:Aquaporins