Purification and functional characterization of aquaporin-8

Kun Liu, Hiroaki Nagase, Chunyi George Huang, Giuseppe Calamita, Peter Agre

Research output: Contribution to journalArticlepeer-review

78 Scopus citations


Background information. Aquaporins (AQPs) are a family of channels permeable to water and some small solutes. In mammals, 13 members (AQP0-AQP12) have been found. AQP8 is widely distributed in many tissues and organs. Previous studies in frog oocytes suggested that AQP8 was permeable to water, urea and ammonium, but no direct characterization had yet been reported. Results. We expressed recombinant rAQP8, hAQP8 a nd mAQP8 (rat, human and mouse AQP8 respectively) in yeast, purified the proteins to homogeneity and reconstituted them into proteoliposomes. Although showing high sequence similarity, AQP8 proteins from the three species had to be purified with different detergents prior to reconstitution. In stopped-flow studies, all three AQP8 proteoliposomes showed water permeability, which was inhibited by mercuric chloride and rescued by 2-mercaptoethanol. rAQP8 and hAQP8 proteoliposomes did not transport glycerol or urea but were permeable to formamide, which was also inhibited by mercuric chloride. In the oocyte transport assay, hAQP8-injected oocytes showed significantly higher [14C]methylammonium uptake than water-injected oocytes. Conclusions. In the present study, we successfully purified rAQP8, hAQP8 and mAQP8 proteins and characterized their biochemical and biophysical properties. All three AQP8 proteins transport water. rAQP8 and hAQP8 are not permeable to urea or glycerol. Moreover, hAQP8 is permeable to ammonium analogues (formamide and methylammonium). Our results suggest that AQP8 may transport ammonium in vivo and physiologically contribute to the acid-base equilibrium.

Original languageEnglish (US)
Pages (from-to)153-161
Number of pages9
JournalBiology of the Cell
Issue number3
StatePublished - Mar 2006


  • Ammonium transport
  • Formamide
  • Methylammonium
  • Proteoliposome
  • Water permeability

ASJC Scopus subject areas

  • Cell Biology


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