Protein stability imposes limits on organism complexity and speed of molecular evolution

Zeldovich, Konstantin B.; Chen, Peiqiu; Shakhnovich, Eugene I.

doi:10.1073/pnas.0705366104

Cited by 245 publications

(354 citation statements)

References 37 publications

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“…Thus, the high stability of the M␤L fold (30) compared with that of serine enzymes (15,16) allows the selection of resistance mutations at the expense of the protein stability. This work also favors the hypothesis put forward by Bloom and coworkers that protein stability promotes evolvability (29) toward other opposite proposals (31).…”

Section: Discussionsupporting

confidence: 67%

Adaptive protein evolution grants organismal fitness by improving catalysis and flexibility

Tomatis

Fabiane

Simona

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

102

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Protein evolution is crucial for organismal adaptation and fitness. This process takes place by shaping a given 3-dimensional fold for its particular biochemical function within the metabolic requirements and constraints of the environment. The complex interplay between sequence, structure, functionality, and stability that gives rise to a particular phenotype has limited the identification of traits acquired through evolution. This is further complicated by the fact that mutations are pleiotropic, and interactions between mutations are not always understood. Antibiotic resistance mediated by ␤-lactamases represents an evolutionary paradigm in which organismal fitness depends on the catalytic efficiency of a single enzyme. Based on this, we have dissected the structural and mechanistic features acquired by an optimized metallo-␤-lactamase (M␤L) obtained by directed evolution. We show that antibiotic resistance mediated by this enzyme is driven by 2 mutations with sign epistasis. One mutation stabilizes a catalytically relevant intermediate by fine tuning the position of 1 metal ion; whereas the other acts by augmenting the protein flexibility. We found that enzyme evolution (and the associated antibiotic resistance) occurred at the expense of the protein stability, revealing that M␤Ls have not exhausted their stability threshold. Our results demonstrate that flexibility is an essential trait that can be acquired during evolution on stable protein scaffolds. Directed evolution aided by a thorough characterization of the selected proteins can be successfully used to predict future evolutionary events and design inhibitors with an evolutionary perspective.antibiotic resistance ͉ enzyme ͉ fitness landscape ͉ metalloproteins ͉ epistasis P rotein evolution is crucial for organismal adaptation and fitness (1-8). This process takes place by shaping a given 3-dimensional fold for its particular biochemical function within the metabolic requirements and constraints of the environment.

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Section: Discussionsupporting

confidence: 67%

Adaptive protein evolution grants organismal fitness by improving catalysis and flexibility

Tomatis

Fabiane

Simona

et al. 2008

Proc. Natl. Acad. Sci. U.S.A.

102

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“…S2A. Likewise, in the absence of selection substitutions are more often destabilizing than stabilizing (binomial test, P < 10 −15 ), as expected from empirical studies on the effects of random mutations (61,72,75,76,80).…”

Section: Resultsmentioning

confidence: 56%

“…Random mutations in a protein-coding sequence typically destabilize the protein structure (26,72,(75)(76)(77)(78)(79)(80). Thus, if protein evolution proceeded solely via random substitutions, without any selection, we would expect a decrease in protein stability over time.…”

Section: Resultsmentioning

confidence: 99%

“…Our model of purifying selection assumes that overstabilizing mutations are as deleterious as destabilizing mutations, so that only the wild-type stability has optimal fitness. However, several studies have shown that overstabilizing mutations can be neutral under stabilizing selection (75,77,80,88). Therefore, we also considered an alternative, semi-Gaussian fitness landscape in which argT sequences more stable than the wild type are just a fit as the wild type (Fig.…”

Section: Robustness Of Simulation Resultsmentioning

confidence: 99%

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Contingency and entrenchment in protein evolution under purifying selection

Shah

McCandlish

Plotkin

2015

Proc. Natl. Acad. Sci. U.S.A.

176

236

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The phenotypic effect of an allele at one genetic site may depend on alleles at other sites, a phenomenon known as epistasis. Epistasis can profoundly influence the process of evolution in populations and shape the patterns of protein divergence across species. Whereas epistasis between adaptive substitutions has been studied extensively, relatively little is known about epistasis under purifying selection. Here we use computational models of thermodynamic stability in a ligand-binding protein to explore the structure of epistasis in simulations of protein sequence evolution. Even though the predicted effects on stability of random mutations are almost completely additive, the mutations that fix under purifying selection are enriched for epistasis. In particular, the mutations that fix are contingent on previous substitutions: Although nearly neutral at their time of fixation, these mutations would be deleterious in the absence of preceding substitutions. Conversely, substitutions under purifying selection are subsequently entrenched by epistasis with later substitutions: They become increasingly deleterious to revert over time. Our results imply that, even under purifying selection, protein sequence evolution is often contingent on history and so it cannot be predicted by the phenotypic effects of mutations assayed in the ancestral background.intragenic epistasis | coevolution | near neutrality | protein stability W hether a heritable mutation is advantageous or deleterious to an organism often depends on the evolutionary history of the population. A mutation that is beneficial at the time of its introduction may confer its beneficial effect only in the presence of other potentiating or permissive mutations (1-9). Thus, the fate of a mutation arising in a population may be contingent on previous mutations (10-13). Conversely, once a mutation has fixed in a population, the mutation becomes part of the genetic background onto which subsequent modifications are introduced. Because the beneficial effects of the subsequent modifications may depend on the focal mutation, as time passes reversion of the focal mutation may become increasingly deleterious, leading to a type of evolutionary conservatism, or entrenchment (14-18).In the context of protein evolution, the effects of contingency and entrenchment are most easily studied by considering a sequence of single amino acid changes (19) that extends both forward and backward in time from some focal substitution. To assess the roles of contingency and entrenchment we can study the degree to which each focal substitution was facilitated by previous substitutions, and the degree to which the focal substitution influences the subsequent course of evolution (Fig. 1A).Dependencies within a sequence of substitutions are closely connected to the concept of epistasis-that is, the idea that the phenotypic effect of a mutation at a particular genetic site may depend on the genetic background in which it arises (20-24). In the absence of epistasis, a mutation has the same effect ...

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“…In addition, increased protein stability is often accompanied by a trade-off in terms of catalytic activity (Somero 1995), although this appears not to be the case for many inferred ancestral enzymes. From an evolutionary perspective, assuming that the increased thermostability of ancestral proteins is not the result of a bias in the in silico reconstruction method (Williams et al 2006; see ''Result and Discussion'' section), the marginal stability of contemporary proteins may imply that an unknown selection pressure has caused the stability of proteins to decrease during evolution, or simply that stability is a neutral trait (Bloom et al 2007;Taverna and Goldstein 2002;Zeldovich et al 2007). For example, as the majority of mutations are destabilising rather than stabilising, it has been argued that proteins only retain high stability if it is evolutionarily beneficial (Taverna and Goldstein 2002).…”

Section: Introductionmentioning

confidence: 99%

Reconstructed Ancestral Enzymes Impose a Fitness Cost upon Modern Bacteria Despite Exhibiting Favourable Biochemical Properties

et al. 2015

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Ancestral sequence reconstruction has been widely used to study historical enzyme evolution, both from biochemical and cellular perspectives. Two properties of reconstructed ancestral proteins/enzymes are commonly reported-high thermostability and high catalytic activity-compared with their contemporaries. Increased protein stability is associated with lower aggregation rates, higher soluble protein abundance and a greater capacity to evolve, and therefore, these proteins could be considered ''superior'' to their contemporary counterparts. In this study, we investigate the relationship between the favourable in vitro biochemical properties of reconstructed ancestral enzymes and the organismal fitness they confer in vivo. We have previously reconstructed several ancestors of the enzyme LeuB, which is essential for leucine biosynthesis. Our initial fitness experiments revealed that overexpression of ANC4, a reconstructed LeuB that exhibits high stability and activity, was only able to partially rescue the growth of a DleuB strain, and that a strain complemented with this enzyme was outcompeted by strains carrying one of its descendants. When we expanded our study to include five reconstructed LeuBs and one contemporary, we found that neither in vitro protein stability nor the catalytic rate was correlated with fitness. Instead, fitness showed a strong, negative correlation with estimated evolutionary age (based on phylogenetic relationships). Our findings suggest that, for reconstructed ancestral enzymes, superior in vitro properties do not translate into organismal fitness in vivo. The molecular basis of the relationship between fitness and the inferred age of ancestral LeuB enzymes is unknown, but may be related to the reconstruction process. We also hypothesise that the ancestral enzymes may be incompatible with the other, contemporary enzymes of the metabolic network.

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Protein stability imposes limits on organism complexity and speed of molecular evolution

Cited by 245 publications

References 37 publications

Adaptive protein evolution grants organismal fitness by improving catalysis and flexibility

Adaptive protein evolution grants organismal fitness by improving catalysis and flexibility

Contingency and entrenchment in protein evolution under purifying selection

Reconstructed Ancestral Enzymes Impose a Fitness Cost upon Modern Bacteria Despite Exhibiting Favourable Biochemical Properties

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