4j8y | pdb_00004j8y

E3_5 DARPin D77S mutant

spinqualitylabelsall models 4j8y, resolution 1.70Å Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image   Print3D Structural highlights 4j8y is a 1 chain structure with sequence from Synthetic construct sequences. Full crystallographic information is available from OCA. For a guided tour on the structure components use FirstGlance. Related: 4j7w, 1mj0 Resources: FirstGlance, OCA, PDBe, RCSB, PDBsum, ProSAT Publication Abstract from PubMed Designed ankyrin repeat proteins (DARPins) are well-established binding molecules based on a highly stable nonantibody scaffold. Building on 13 crystal structures of DARPin-target complexes and stability measurements of DARPin mutants, we have generated a new DARPin library containing an extended randomized surface. To counteract the enrichment of unspecific hydrophobic binders during selections against difficult targets containing hydrophobic surfaces such as membrane proteins, the frequency of apolar residues at diversified positions was drastically reduced and substituted by an increased number of tyrosines. Ribosome display selections against two human caspases and membrane transporter AcrB yielded highly enriched pools of unique and strong DARPin binders which were mainly monomeric. We noted a prominent enrichment of tryptophan residues during binder selections. A crystal structure of a representative of this library in complex with caspase-7 visualizes the key roles of both tryptophans and tyrosines in providing target contacts. These aromatic and polar side chains thus substitute the apolar residues valine, leucine, isoleucine, methionine, and phenylalanine of the original DARPins. Our work describes biophysical and structural analyses required to extend existing binder scaffolds and simplifies an existing protocol for the assembly of highly diverse synthetic binder libraries. Design, construction, and characterization of a second-generation DARPin library with reduced hydrophobicity.,Seeger MA, Zbinden R, Flutsch A, Gutte PG, Engeler S, Roschitzki-Voser H, Grutter MG Protein Sci. 2013 Jul 19. doi: 10.1002/pro.2312. PMID:23868333 From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.

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