Large‐scale purification of functional recombinant human aquaporin‐2

FEBS Letters

Koenderink, Jan B.¹  Deen, Peter M.T.²  Hasler, Lorenz³  Klaassen, Corné H.W.¹  Werten, Paul J.L.²  de Grip, Willem J.¹  Engel, Andreas³ 

[1] Department of Biochemistry, University of Nijmegen, Nijmegen, The Netherlands;Department of Cell Physiology, University of Nijmegen, Nijmegen, The Netherlands;ME Müller Institut, Biozentrum, Universität Basel, Basel, Switzerland

关键词: Baculovirus;    Insect cell;    Expression;    Stopped-flow analysis;    Structural analysis;   

DOI  :  10.1016/S0014-5793(01)02703-X

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

The homotetrameric aquaporin-2 (AQP2) water channel is essential for the concentration of urine and of critical importance in diseases with water dysregulation, such as nephrogenic diabetes insipidus, congestive heart failure, liver cirrhosis and pre-eclampsia. The structure of human AQP2 is a prerequisite for understanding its function and for designing specific blockers. To obtain sufficient amounts of AQP2 for structural analyses, we have expressed recombinant his-tagged human AQP2 (HT-AQP2) in the baculovirus/insect cell system. Using the protocols outlined in this study, 0.5 mg of pure HT-AQP2 could be obtained per liter of bioreactor culture. HT-AQP2 had retained its homotetrameric structure and exhibited a single channel water permeability of 0.93±0.03×10⁻¹³ cm³/s, similar to that of other AQPs. Thus, the baculovirus/insect cell system allows large-scale expression of functional recombinant human AQP2 that is suitable for structural studies.

【 授权许可】

Unknown