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Tethering Polypeptides Through Bifunctional Peg Cross-Linking Agents To Probe Protein Function: Application To Atp Synthase., Daniel J Cipriano, Stanley D Dunn

Biochemistry Publications

Chemical crosslinking mediated by short bifunctional reagents has been widely used for determining physical relationships among polypeptides in multisubunit proteins, but less often for functional studies. Here we introduce the approach of tethering polypeptides by using bifunctional reagents containing a lengthy, flexible PEG linker as a form of crosslinking especially suited to functional analyses. The rotary molecular motor ATP synthase was used as a model subject. Single cysteine residues were introduced into selected positions of ATP synthase epsilon subunit, a component of the rotor subcomplex of the enzyme, and the unrelated maltose binding protein (MBP), then the two purified recombinant …