Structural Basis of Furan–Amino Acid Recognition by a Polyspecific Aminoacyl‐tRNA‐Synthetase and its Genetic Encoding in Human Cells. Issue 12 (15th April 2014) - Schmidt, Moritz J.
Abstract The site‐selective introduction of photo‐crosslinking groups into proteins enables the discovery and mapping of weak and/or transient protein interactions with high spatiotemporal resolution, both in vitro and in vivo. We report the genetic encoding of a furan‐based, photo‐crosslinking amino acid in human cells; it can be activated with red light, thus offering high penetration depths in biological samples. This is achieved by activation of the amino acid and charging to its cognate tRNA by a pyrrolysyl‐tRNA‐synthetase (PylRS) mutant with broad polyspecificity. To gain insights into the recognition of this amino acid and to provide a rationale for its polyspecificity, we solved three crystal structures of the PylRS mutant: in its apo‐form, in complex with adenosine 5′‐(β, γ‐imido)triphosphate (AMP‐PNP) and in complex with the AMP ester of the furan amino acid. These structures provide clues for the observed polyspecificity and represent a promising starting point for the engineering of PylRS mutants with further increased substrate scope.