Alpha repeat proteins ( αRep) as expression and crystallization helpers

Publication date: Available online 16 August 2017 Source:Journal of Structural Biology Author(s): Anne Chevrel, Agnes Mesneau, Dyana Sanchez, Louisa Celma, Sophie Quevillon-Cheruel, Andrea Cavagnino, Sylvie Nessler, Ines Li de la Sierra-Gallay, Herman van Tilbeurgh, Philippe Minard, Marie Valerio-Lepiniec, Agathe Urvoas We have previously described a highly diverse library of artificial repeat proteins based on thermostable HEAT-like repeats, named αRep. αReps binding specifically to proteins difficult to crystallize have been selected and in several examples, they made possible the crystallization of these proteins. To further simplify the production and crystallization experiments we have explored the production of chimeric proteins corresponding to covalent association between the targets and their specific binders strengthened by a linker. Although chimeric proteins with expression partners are classically used to enhance expression, these fusions cannot usually be used for crystallization. With specific expression partners like a cognate αRep this is no longer true, and chimeric proteins can be expressed purified and crystallized. αRep selection by phage display suppose that at least a small amount of the target protein should be produced to be used as a bait for selection and this might, in some cases, be difficult. We have therefore transferred the αRep library in a new construction adapted to selection by protein complementation assay (PCA). This ne...
Source: Journal of Structural Biology - Category: Biology Source Type: research