Predicting affinity- and specificity-enhancing mutations at protein-protein interfaces

Oz Sharabi, Jason Shirian, Julia M. Shifman*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

18 Scopus citations


Manipulations of PPIs (protein-protein interactions) are important for many biological applications such as synthetic biology and drug design. Combinatorial methods havebeen traditionally used for such manipulations, failing, however, to explain the effects achieved. We developed a computational method for prediction of changes in free energy of binding due tomutation that bring about deeper understanding of the molecular forces underlying binding interactions. Our method could be used for computational scanning of binding interfaces and subsequent analysis of the interfacial sequence optimality. The computational method was validated in two biological systems. Computational saturated mutagenesis of a high-affinity complex between an enzyme AChE (acetylcholinesterase) and a snake toxin Fas (fasciculin) revealed the optimal nature of this interface withonly a few predicted affinity-enhancing mutations. Binding measurements confirmed high optimality of this interface and identified a few mutations that could further improve interaction fitness. Computational interface scanning of a medium-affinity complex between TIMP-2 (tissue inhibitor of metalloproteinases-2) and MMP (matrix metalloproteinase) 14 revealed a non-optimal nature of the binding interface with multiple mutations predicted to stabilize the complex. Experimental results corroborated our computational predictions, identifying a large number of mutations that improve the binding affinity for this interaction and some mutations that enhance binding specificity. Overall, our computational protocol greatly facilitates the discovery of affinity- and specificity-enhancing mutations and thus could be applied for design of potent and highly specific inhibitors of any PPI.

Original languageAmerican English
Pages (from-to)1166-1169
Number of pages4
JournalBiochemical Society Transactions
Issue number5
StatePublished - Oct 2013


  • Binding affinity
  • Binding landscape
  • Protein design
  • Protein-protein interaction
  • Saturated mutagenesis


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