Nature 459, 668-673 (4 June 2009) | doi:10.1038/nature08009; Received 4 June 2008; Accepted 20 March 2009

Chaperonin overexpression promotes genetic variation and enzyme evolution

Nobuhiko Tokuriki1 & Dan S. Tawfik1

  1. Department of Biological Chemistry, Weizmann Institute of Science, Rehovot 76100, Israel

Correspondence to: Dan S. Tawfik1 Correspondence and requests for materials should be addressed to D.S.T. (Email:


Most protein mutations, and mutations that alter protein functions in particular, undermine stability and are therefore deleterious. Chaperones, or heat-shock proteins, are often implicated in buffering mutations, and could thus facilitate the acquisition of neutral genetic diversity and the rate of adaptation. We examined the ability of the Escherichia coli GroEL/GroES chaperonins to buffer destabilizing and adaptive mutations. Here we show that mutational drifts performed in vitro with four different enzymes indicated that GroEL/GroES overexpression doubled the number of accumulating mutations, and promoted the folding of enzyme variants carrying mutations in the protein core and/or mutations with higher destabilizing effects (destabilization energies of >3.5 kcal mol- 1, on average, versus approx1 kcal mol- 1 in the absence of GroEL/GroES). The divergence of modified enzymatic specificity occurred much faster under GroEL/GroES overexpression, in terms of the number of adapted variants (greater than or equal to2-fold) and their improved specificity and activity (greater than or equal to10-fold). These results indicate that protein stability is a major constraint in protein evolution, and buffering mechanisms such as chaperonins are key in alleviating this constraint.


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