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Updated: Jun 1, 2026

Determining Membrane Protein Topology Using Fluorescence Protease Protection (FPP)
Published on: April 20, 2015
Flanking residues help determine whether a hydrophobic segment adopts a monotopic or bitopic topology in the
Morten H H Nørholm1, Yulia V Shulga, Satoko Aoki
1Center for Biomembrane Research, Department of Biochemistry and Biophysics, Stockholm University, SE-106 91 Stockholm, Sweden.
Abstract:
Proteins interacting with membranes via a single hydrophobic segment can be classified as either monotopic or bitopic. Here, we probe the topology of a membrane-attached enzyme, the ε isoform of human diacylglycerol kinase (DGKε), when inserted into rough microsomes and compare it with the monotopic membrane protein mouse caveolin-1. In contrast to previous findings, the N-terminal hydrophobic stretch in DGKε attains a bitopic rather than a monotopic topology in our experimental system. In addition, we find that charged flanking residues as well as proline residues embedded in the hydrophobic segment are important determinants of monotopic versus bitopic topology.
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