1vpo
From Proteopedia
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Crystal Structure Analysis of the Anti-testosterone Fab in Complex with Testosterone
Overview
A highly selective, high affinity recombinant anti-testosterone Fab, fragment has been generated by stepwise optimization of the, complementarity-determining regions (CDRs) by random mutagenesis and phage, display selection of a monoclonal antibody (3-C(4)F(5)). The best mutant, (77 Fab) was obtained by evaluating the additivity effects of different, independently selected CDR mutations. The 77 Fab contains 20 mutations and, has about 40-fold increased affinity (K(d) = 3 x 10(-10) m) when compared, with the wild-type (3-C(4)F(5)) Fab. To obtain structural insight into, factors, which are needed to improve binding properties, we have, determined the crystal structures of the mutant 77 Fab fragment with (2.15, A) and without testosterone (2.10 A) and compared these with previously, determined wild-type structures. The overall testosterone binding of the, 77 Fab is similar to that of the wild-type. The improved affinity and, specificity of the 77 Fab fragment are due to more comprehensive packing, of the testosterone with the protein, which is the result of small, structural changes within the variable domains. Only one important binding, site residue Glu-95 of the heavy chain CDR3 is mutated to alanine in the, 77 Fab fragment. This mutation, originally selected from the phage library, based on improved specificity, provides more free space for the, testosterone D-ring. The light chain CDR1 of 77 Fab containing eight, mutations has the most significant effect on the improved affinity, although it has no direct contact with the testosterone. The mutations of, CDR-L1 cause a rearrangement in its conformation, leading to an overall, fine reshaping of the binding site.
About this Structure
1VPO is a Protein complex structure of sequences from Mus musculus with TES as ligand. This structure superseeds the now removed PDB entry 1L7S. Full crystallographic information is available from OCA.
Reference
Crystal structure of an in vitro affinity- and specificity-matured anti-testosterone Fab in complex with testosterone. Improved affinity results from small structural changes within the variable domains., Valjakka J, Hemminki A, Niemi S, Soderlund H, Takkinen K, Rouvinen J, J Biol Chem. 2002 Nov 15;277(46):44021-7. Epub 2002 Aug 23. PMID:12196551
Page seeded by OCA on Sat Nov 24 22:58:24 2007
