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Technology Topics Modeling

Metal mates

SBKB [doi:10.1038/sbkb.2011.63]
Technical Highlight - February 2012
Short description: Experimental validation of a designed metal-mediated dimer highlights the potential of computational methods.

Comparison of the MID1 model (green) and crystal structure (blue). Spheres indicate zinc ions. Figure courtesy of Brian Kuhlman.

The diversity of protein-protein interactions presents a significant challenge to the rational design of novel interfaces. While directed evolution is a powerful method, computational design allows specification of new interfaces in desired locations and orientations. This approach does, however, have drawbacks associated with the enormous conformational search space and requires accurate modeling of atomic details at the interface.

Kuhlman and colleagues (PSI NESG) have used known coordination spheres of metals to design and experimentally validate the structure of a metal-mediated symmetric homodimer. The authors used RosettaMatch to select 42,000 two-residue zinc binding matches on 600 known monomeric protein scaffolds. The matches were used to combinatorially generate 500,000 dimeric coordinate sets with no backbone clashes and acceptable tetrahedral coordination geometry for the zinc ions. These starting structures were input into a design step that iteratively optimized both backbone geometry and sequence while minimizing the number of mutations. Finally, the models were evaluated by Rosetta analysis of the binding interfaces and zinc positions, allowing the authors to select eight designs for experimental validation.

Although seven of the eight designs expressed poorly or formed higher-order oligomers, metal interface design 1 (MID1) dimerized as expected. The authors used fluorescence polarization to measure an impressive 200-fold increase in binding affinity upon zinc coordination. Although conformational heterogeneity precluded the use of NMR, the authors were able to determine the crystal structures of wild-type and mutant forms of MID1. Structures of the MID1-apo dimer do not resemble the computational design. However, the authors observed a reorientation of the dimer interface in the zinc-coordinated structure in which the zinc atoms and interface residues are very close to the designed positions in a nearly symmetric dimer. The structure further shows that the zinc ions are coordinated by three histidines compared to the four designed. The authors repaired the four-residue zinc coordination by mutation or cobalt substitution while noting that four-histidine coordination is a very rare natural occurrence. The results contribute to our understanding of protein complexes and have practical implications for therapeutic protein design and synthetic biology.

Michael A. Durney


  1. B. S. Der et al. Metal-mediated affinity and orientation specificity in a computationally designed protein homodimer.
    J. Am. Chem. Soc. (17 November 2011). doi:10.1021/ja208015j

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