The mechanical stability of ubiquitin is linkage dependent

Abstract

Ubiquitin chains are formed through the action of a set of enzymes that covalently link ubiquitin either through peptide bonds or through isopeptide bonds between their C terminus and any of four lysine residues. These naturally occurring polyproteins allow one to study the mechanical stability of a protein, when force is applied through different linkages. Here we used single-molecule force spectroscopy techniques to examine the mechanical stability of N-C–linked and Lys48-C–linked ubiquitin chains. We combined these experiments with steered molecular dynamics (SMD) simulations and found that the mechanical stability and unfolding pathway of ubiquitin strongly depend on the linkage through which the mechanical force is applied to the protein. Hence, a protein that is otherwise very stable may be easily unfolded by a relatively weak mechanical force applied through the right linkage. This may be a widespread mechanism in biological systems.

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Figure 1: Mechanical properties of N-C polyubiquitin.
Figure 2: The linkage between domains markedly affects the mechanical properties of ubiquitin.
Figure 3: The mechanical unfolding forces of ubiquitin chains depends on the rate at which they are pulled.
Figure 4: The linkage-dependent properties of ubiquitin reproduced in silico using steered molecular dynamics.

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Acknowledgements

We thank J. Vuust (Statens Serum Institut, Denmark) for his gift of the human Ubi9 cDNA clone. We thank K. Schulten (University of Illinois, Urbana-Champaign) for his support in building and operating our PC-based Beowulf cluster for molecular dynamics simulations. This work was funded by grants from the National Institutes of Health to J.M.F.

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Correspondence to Julio M Fernandez.

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