UNIT 5.2 Determining the Topology of an Integral Membrane Protein
Published Online: 1 MAY 2001
Copyright © 2003 by John Wiley and Sons, Inc.
Lab Protocol Title
Current Protocols in Cell Biology
How to Cite
Green, N., Fang, H., Kalies, K.-U. and Canfield, V. 2001. Determining the Topology of an Integral Membrane Protein. Current Protocols in Cell Biology. 00:5.2:5.2.1–5.2.27.
- Published Online: 1 MAY 2001
- Published Print: JAN 2000
A variety of methods have been developed for assigning the aqueous domains of integral membrane proteins to either side of a biological membrane. Once the sequence of a protein is known from its DNA sequence it is possible to study the topology of the protein. This unit provides protocols in which the water-soluble domains can be tested for their accessibility to reagents added to membranes with a defined orientation. Tagging of hydrophilic regions of the protein with different epitopes and probing of their orientation with respect to the membrane is also described. Finally, a procedure for fusion of a reporter enzyme to truncated fragments of the protein is provided. The fusion protein is used as a sensor of sequence disposition relative to the membrane.