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The spatial architecture of protein function and adaptation


Statistical analysis of protein evolution suggests a design for natural proteins in which sparse networks of coevolving amino acids (termed sectors) comprise the essence of three-dimensional structure and function1,2,3,4,5. However, proteins are also subject to pressures deriving from the dynamics of the evolutionary process itself—the ability to tolerate mutation and to be adaptive to changing selection pressures6,7,8,9,10. To understand the relationship of the sector architecture to these properties, we developed a high-throughput quantitative method for a comprehensive single-mutation study in which every position is substituted individually to every other amino acid. Using a PDZ domain (PSD95pdz3) model system, we show that sector positions are functionally sensitive to mutation, whereas non-sector positions are more tolerant to substitution. In addition, we find that adaptation to a new binding specificity initiates exclusively through variation within sector residues. A combination of just two sector mutations located near and away from the ligand-binding site suffices to switch the binding specificity of PSD95pdz3 quantitatively towards a class-switching ligand. The localization of functional constraint and adaptive variation within the sector has important implications for understanding and engineering proteins.

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Figure 1: Sector architecture in the PDZ domain family.
Figure 2: Complete single mutagenesis in PSD95pdz3.
Figure 3: The relationship of mutational sensitivity of positions to the protein sector.
Figure 4: Adaptation through sector variation.


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We thank W. Russ, P. Mishra and other members of the Ranganathan laboratory for contributions to this work, W. Wakeland and C. Liang for assistance with Solexa sequencing, E. Curry and A. Mobley for assistance with flow cytometry, and M. Elowitz for providing the pZ plasmids. We acknowledge support from the University of Texas Southwestern Graduate School and Pharmacology Training Grant (R. N. M.), the Helen Hay Whitney Fellowship program (F.J.P.) and support from the National Institutes of Health (R01EY018720-05, R.R.), The Robert A. Welch Foundation (I-1366, R.R.) and the Green Center for Systems Biology (R.R.).

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Authors and Affiliations



R.N.M. and R.R. developed the research plan and experimental strategy. R.N.M. built the B2H assay, collected the data and wrote and executed the code for analysis of the B2H and sequencing data. F.J.P. and W.S.G. improved the dynamic range of the B2H assay. A.R. carried out the mutational analysis in Fig. 4 e, f. R.N.M. and R.R. analysed the data and wrote the paper.

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Correspondence to Rama Ranganathan.

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The authors declare no competing financial interests.

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McLaughlin Jr, R., Poelwijk, F., Raman, A. et al. The spatial architecture of protein function and adaptation. Nature 491, 138–142 (2012).

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