Characterisation of the Major Intrinsic Protein (MIP) from Bovine Lens Fibre Membranes by Electron Microscopy and Hydrodynamics

Authors: König N.¹; Zampighi G.A.²; Butler P.J.G.¹

Source: Journal of Molecular Biology, Volume 265, Number 5, January 1997 , pp. 590-602(13)

Publisher: Academic Press

Abstract:

The major intrinsic protein (MIP) from bovine lens fibre membranes has been purified from unstripped membranes using a single ion-exchange chromatography step (MonoS) in the non-ionic detergent octyl-- d -glucopyranoside (OG). SDS-PAGE has confirmed the purity of the preparation and thin-layer chromatographic analysis has shown that the protein is virtually lipid-free. To establish a stable and monodisperse protein sample, we exchanged OG with decyl-- d -maltopyranoside (DeM), another non-ionic detergent, by gel-filtration column chromatography. We conclude that the resulting protein/detergent complex is composed of four copies of MIP (a tetramer) and a detergent micelle. This conclusion is based on: (1) measurement of the weight-average molecular mass (M_w,app) of the protein moiety in the protein/detergent complex by sedimentation equilibrium; (2) measurement of the apparent molecular mass of the complexes formed by MIP in OG, in DeM, in dodecyl-- d -maltopyranoside (DoM) and in sodium dodecylsulphate (SDS) by gel filtration; (3) measurement of the apparent molecular mass of pure detergent micelles; (4) measurement of the predicted change in the molecular mass of the MIP/DeM complex after partial enzymatic proteolysis; and (5) measurement of the size and shape of the MIP/detergent complex by electron microscopy and single-particle analysis. Therefore, the tetragonal arrangement of MIP observed in both plasma membranes and junctional membranes in lens fibre cells is maintained in solution with non-ionic detergents. Copyright 1997 Academic Press

Keywords: MIP; major intrinsic protein; lens fibre cells; purification; tetramer

Language: English

Document Type: Research article

Affiliations: 1: MRC Laboratory of Molecular Biology, Hills Road, Cambridge, CB2 2QH, UK 2: UCLA School of Medicine, Department of Neurobiology, Los Angeles, CA 90024, USA

Publication date: 1997-01-01

• In this: publication
• By this: publisher
• In this Subject: Biology ,  Microbiology
• By this author: König N. ;  Zampighi G.A. ;  Butler P.J.G.

Website © Publishing Technology. Article copyright remains with the publisher, society or author(s) as specified within the article.

• Terms and conditions
• Privacy policy
• Information for advertisers