Achieving photo-control of protein conformation and activity: producing a photo-controlled leucine zipper

We have recently developed a technique that has great potential in producing proteins with photo-control of conformation and consequently activity (J. R. Kumita, O. S. Smart and G. A. Woolley, Proc. Natl. Acad. Sci. U. S. A., 2000, 97, 3803 3808). The method is based on incorporating two cysteine residues into the sequence of a polypeptide. An azobenzene derivative is subsequently used to produce an intramolecular cross-link between the cysteine sulfhydryl groups. In previous work photo-isomerisation of the azobenzene moiety has been used to control the helicity of a monomeric peptide. In the experiments described here this method has been applied to the coiled coil leucine zipper peptide GCN4-p1. The aim was to produce a variant of GCN4-p1 whose helicity and consequently dimerisation is under direct photo-control. We have produced a modified GCN4-p1 incorporating two cysteine residues. The mutations introduced are shown to interfere with the ability of the uncross-linked peptide to form a coiled coil. After the peptide was cross-linked with the azobenzene derivative more normal coiled-coil behaviour was restored. Irradiation of the peptide producing a conformational change in the azobenzene cross-linker was accompanied by an increase in the helicity of the peptide. The work presented here highlights the potential of the use of photo-isomerisable cross-linkers to control protein activity through induced conformational change. In addition, the methodology has the potential to provide a fast trigger for the initiation of protein conformational changes.