4JB8

Caspase-7 in Complex with DARPin C7_16

Summary for 4JB8

• Entry DOI: 10.2210/pdb4jb8/pdb
• Related: 3IBC 4J92
• Descriptor: Caspase-7 subunit p20, Caspase-7 subunit p11, DARPin C7_16, ... (4 entities in total)
• Functional Keywords: complex structure, selected darpin c7_16, hydrolase-de novo protein complex, hydrolase/de novo protein
• Biological source: Homo sapiens (human); More
• Cellular location: Cytoplasm: P55210 P55210
• Total number of polymer chains: 3
• Total formula weight: 50378.71

Authors

Fluetsch, A.,Seeger, M.A.,Gruetter, M.G. (deposition date: 2013-02-19, release date: 2013-08-21, Last modification date: 2023-09-20)

Primary citation

Seeger, M.A.,Zbinden, R.,Flutsch, A.,Gutte, P.G.,Engeler, S.,Roschitzki-Voser, H.,Grutter, M.G.
Design, construction, and characterization of a second-generation DARPin library with reduced hydrophobicity.
Protein Sci., 22:1239-1257, 2013

PubMed Abstract: 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.

PubMed: 23868333
DOI: 10.1002/pro.2312

Experimental method

X-RAY DIFFRACTION (1.7 Å)