Action‐at‐a‐distance interactions enhance protein binding affinity

Abstract

The identification of protein mutations that enhance binding affinity may be achieved by computational or experimental means, or by a combination of the two. Sources of affinity enhancement may include improvements to the net balance of binding interactions of residues forming intermolecular contacts at the binding interface, such as packing and hydrogen-bonding interactions. Here we identify noncontacting residues that make substantial contributions to binding affinity and that also provide opportunities for mutations that increase binding affinity of the TEM1 ␤-lactamase (TEM1) to the ␤-lactamase inhibitor protein (BLIP). A region of BLIP not on the direct TEM1-binding surface was identified for which changes in net charge result in particularly large increases in computed binding affinity. Some mutations to the region have previously been characterized, and our results are in good correspondence with this results of that study. In addition, we propose novel mutations to BLIP that were computed to improve binding significantly without contacting TEM1 directly. This class of noncontacting electrostatic interactions could have general utility in the design and tuning of binding interactions.Keywords: continuum electrostatics; electrostatic complementarity; protein binding; protein-protein interactions; protein designThe field of protein design has made substantial advances over the last 20 years, based largely on phrasing the appropriate inverse problem and developing methods capable of addressing inverse design (Drexler 1981;Pabo 1983). Much current protein design work involves the construction of stabilizing protein side-chain arrangements by methods such as dead-end elimination (Desmet et al. 1992;Goldstein 1994;Lasters et al. 1995;Gordon andMayo 1998, 1999;Leach and Lemon 1998;Mendes et al. 1999;De Maeyer et al. 2000;Looger and Hellinga 2001), self-consistent meanfield theory (Koehl and Delarue 1994; Koehl and Levitt 1999a,b;Kono and Saven 2001), simulated annealing (Lee and Subbiah 1991;Hellinga and Richards 1994;Shenkin et al. 1996;Jiang et al. 1997Jiang et al. , 2000, genetic algorithms (Tufféry et al. 1991(Tufféry et al. , 1993(Tufféry et al. , 1997Jones 1994;Desjarlais and Handel 1995), and combinatorial search (Tufféry et al. 1991(Tufféry et al. , 1993(Tufféry et al. , 1997. That is, successful design has been achieved by consideration of detailed atomic interactions and their effect on packing geometry and energetics (Dahiyat and Mayo 1997;Harbury et al. 1998;Calhoun et al. 2003;Kuhlman et al. 2003). The design of protein binding interfaces may be achieved by a similar overall approach, although the additional requirement to treat solvation and electrostatic interactions adds a further layer of complexity (Lee and Tidor 2001a).An alternative strategy that does not demand the same detailed packing of side chains into an exquisite three-dimensional jigsaw puzzle may be desirable in many cases. One such method involves the enhancement of affinity through relatively long-range electrostatic ef...

Fine Mapping of the Sequence Requirements for Binding of β-Lactamase Inhibitory Protein (BLIP) to TEM-1 β-Lactamase Using a Genetic Screen for BLIP Function

“…In addition, high-resolution co-crystal structures of BLIP in complex with TEM-1 and SHV-1 β-lactamase are available 18; 19. The BLIP-β-lactamase interface has been extensively studied using structural, computational and biochemical approaches 18; 19; 20; 21; 22; 23; 24; 25. Extensive site-directed mutagenesis experiments by Schreiber and colleagues suggest the interface consists of modules of TEM-1 and BLIP residues where cooperative interactions occur between residues within modules and additive interactions occur among residues in different modules 26; 27.…”

Insights into Positive and Negative Requirements for Protein–Protein Interactions by Crystallographic Analysis of the β-Lactamase Inhibitory Proteins BLIP, BLIP-I, and BLP

“…The originally identified BLIP (from Streptomyces clavuligerus) 9 has become a model for a broad range of structural, mutagenic, biophysical, and computational studies. [10][11][12][13][14][15][16][17][18][19][20][21] BLIP has two close probable homologues: BLIP-I (from Streptomyces exfoliatus) and β-lactamase-inhibitoryprotein-like protein (BLP; S. clavuligerus). No other related protein † is found in any Streptomyces genome that has been sequenced fully ‡ or partially §.…”

Computational design of antibody-affinity improvement beyond in vivo maturation

“…In a second mechanism, the addition of a charged residue, net charge is changed to increase electrostatic interaction, often at the periphery of the antibody-antigen interface where desolvation is minimal. Unlike previous work using electrostatics to guide design 25,26 , these methods explicitly model the mutation, calculate a binding free energy relative to wild type, include positions that are partially or fully buried upon binding, and avoid opportunities where the mutation is predicted to destabilize the mutant protein.The modeled structures result from optimization with the full energy function, yet the van der Waals and nonpolar solvation energies are then discarded to predict improvement based on only the net electrostatic term. A potential problem is that different/wrong van der Waals parameters would produce altered equilibrium distances and hence altered electrostatic energies, particularly for short-range interactions.…”

“…In a second mechanism, the addition of a charged residue, net charge is changed to increase electrostatic interaction, often at the periphery of the antibody-antigen interface where desolvation is minimal. Unlike previous work using electrostatics to guide design 25,26 , these methods explicitly model the mutation, calculate a binding free energy relative to wild type, include positions that are partially or fully buried upon binding, and avoid opportunities where the mutation is predicted to destabilize the mutant protein.…”