Genetic Selection for Critical Residues in Ribonucleases

Smith, Bryan D.; Raines, Ronald T.

doi:10.1016/j.jmb.2006.07.020

Cited by 32 publications

(58 citation statements)

References 129 publications

(202 reference statements)

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“…Other cellular biomolecules can serve to regulate the cellular redox environment (79), but no other cellular inhibitors of ribonucleases are known (26). Consequently, RI needs to maintain its high affinity for ribonucleases so as to protect cellular RNA, even under severely oxidizing conditions (12,13,15).…”

Section: Discussionmentioning

confidence: 99%

“…Nonetheless, the co-evolution of certain non-obligate protein-protein interactions, such as particular receptor-ligand and enzyme-inhibitor interactions, can be essential for cell survival (11). To understand the co-evolution of such a non-obligate pair, we have investigated the complex formed between the cytosolic ribonuclease inhibitor protein (RI 1 ) and secretory pancreatictype ribonucleases, as failure to inhibit the enzymatic activity of an invading ribonuclease can lead to cell death (12)(13)(14)(15) RIs (25,26), which binds to some members of the ribonuclease A superfamily with affinities in the femtomolar range (27,28). RI is able to exert this affinity using its large concave surface area (28-30), even though the sequence identity among superfamily members is <30% (26,28).…”

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confidence: 99%

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Intraspecies Regulation of Ribonucleolytic Activity

2007

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The evolutionary rate of proteins involved in obligate protein-protein interactions is slower and the degree of co-evolution higher than that for non-obligate protein-protein interactions. The coevolution of the proteins involved in certain non-obligate interactions is, however, essential to cell survival. To gain insight into the co-evolution of one such non-obligate protein pair, the cytosolic ribonuclease inhibitor (RI) proteins and secretory pancreatic-type ribonucleases from cow (Bos taurus) and human (Homo sapiens) were produced in Escherichia coli and purified, and their physicochemical properties were analyzed. The two intraspecies complexes were found to be extremely tight (bovine K d = 0.69 fM; human K d = 0.34 fM). Human RI binds to its cognate ribonuclease (RNase 1) with 100-fold greater affinity than to the bovine homologue (RNase A). In contrast, bovine RI binds to RNase 1 and RNase A with nearly equal affinity. This broader specificity is consistent with there being more pancreatic-type ribonucleases in cows (20) than humans (13). Human RI (32 cysteine residues) also has 4-fold less resistance to oxidation by hydrogen peroxide than does bovine RI (29 cysteine residues). This decreased oxidative stability of human RI, which is caused largely by Cys74, implies a larger role for human RI as an antioxidant. The conformational and oxidative stabilities of both RIs increase upon complex formation with ribonucleases. Thus, RI has evolved to maintain its inhibition of invading ribonucleases, even when confronted with extreme environmental stress. That role appears to take precedence over its role in mediating oxidative damage.The discovery of extensive protein-protein interaction networks has informed investigations of protein evolution (1-3). For example, the rate of evolution within a protein-protein interaction network, has been found to depend on the number of binding partners, the concentration of each protein, and the evolutionary age of the proteins (4-10). These relationships extend to subclasses of interactions, as obligate protein-protein interactions, which are defined as interactions necessary for the stability of an individual protein (11), evolve at a slower rate and show a greater degree of co-evolution than do non-obligate interactions, which are interactions between proteins that can remain stable independently (7). Nonetheless, the co-evolution of certain non-obligate protein-protein interactions, such as particular receptor-ligand and enzyme-inhibitor interactions, can be essential for cell survival (11). To understand the co-evolution of such a non-obligate pair, we have investigated the complex formed between the cytosolic ribonuclease inhibitor protein (RI 1 ) and secretory pancreatictype ribonucleases, as failure to inhibit the enzymatic activity of an invading ribonuclease can lead to cell death (12)(13)(14)(15) RIs (25,26), which binds to some members of the ribonuclease A superfamily with affinities in the femtomolar range (27,28). RI is able to exert this affinity using...

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Section: Discussionmentioning

confidence: 99%

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confidence: 99%

Intraspecies Regulation of Ribonucleolytic Activity

2007

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“…Mutagenesis experiments have shown that different positions in proteins have widely differing tolerances to amino acid substitutions (Reidhaar-Olson and Sauer 1988;Bowie et al 1990;Lau and Dill 1990;Guo et al 2004;Campbell-Valois et al 2005;Smith and Raines 2006). On average, however, mutations introduced at solvent-exposed sites are less likely to disrupt protein structure and function than mutations introduced at buried sites.…”

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The Relationship Between Relative Solvent Accessibility and Evolutionary Rate in Protein Evolution

Ramsey

Scherrer

Zhou³

et al. 2011

Genetics

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Recent work with Saccharomyces cerevisiae shows a linear relationship between the evolutionary rate of sites and the relative solvent accessibility (RSA) of the corresponding residues in the folded protein. Here, we aim to develop a mathematical model that can reproduce this linear relationship. We first demonstrate that two models that both seem reasonable choices (a simple model in which selection strength correlates with RSA and a more complex model based on RSA-dependent amino acid distributions) fail to reproduce the observed relationship. We then develop a model on the basis of observed site-specific amino acid distributions and show that this model behaves appropriately. We conclude that evolutionary rates are directly linked to the distribution of amino acids at individual sites. Because of this link, any future insight into the biophysical mechanisms that determine amino acid distributions will improve our understanding of evolutionary rates.T HE requirement for successful and efficient protein folding imposes significant biophysical constraints on coding sequences. These constraints shape how sequences evolve. Mutations that interfere with correct folding will generally be removed by purifying selection. Likewise, mutations that do not interfere with folding are often neutral, or nearly so, and accumulate over time. As a consequence of this interaction between protein biophysics and molecular evolution, signatures of protein structure can be found in the divergence

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“…Loss in binding energy (16 kJ mol À1 ) by a double mutation was comparable to RI binding to S-protein alone. 58 Smith and Raines 59 have developed a creative expression system in an E. coli strain in which the reductive intracellular environment has been eliminated to allow nascent RNase to form its disulfide bonds and generate enzymatic activity. Mutations within this E. coli strain allowed the interface between RNase and RI to be mapped.…”

Section: Intracellular Inhibitors Of Rnasementioning

confidence: 99%

Back to the future: Ribonuclease A

2008

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Pancreatic ribonuclease A (EC 3.1.27.5, RNase) is, perhaps, the best‐studied enzyme of the 20th century. It was isolated by René Dubos, crystallized by Moses Kunitz, sequenced by Stanford Moore and William Stein, and synthesized in the laboratory of Bruce Merrifield, all at the Rockefeller Institute/University. It has proven to be an excellent model system for many different types of experiments, both as an enzyme and as a well‐characterized protein for biophysical studies. Of major significance was the demonstration by Chris Anfinsen at NIH that the primary sequence of RNase encoded the three‐dimensional structure of the enzyme. Many other prominent protein chemists/enzymologists have utilized RNase as a dominant theme in their research. In this review, the history of RNase and its offspring, RNase S (S‐protein/S‐peptide), will be considered, especially the work in the Merrifield group, as a preface to preliminary data and proposed experiments addressing topics of current interest. These include entropy–enthalpy compensation, entropy of ligand binding, the impact of protein modification on thermal stability, and the role of protein dynamics in enzyme action. In continuing to use RNase as a prototypical enzyme, we stand on the shoulders of the giants of protein chemistry to survey the future. © 2007 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 90: 259–277, 2008.This article was originally published online as an accepted preprint. The “Published Online” date corresponds to the preprint version. You can request a copy of the preprint by emailing the Biopolymers editorial office at biopolymers@wiley.com

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Genetic Selection for Critical Residues in Ribonucleases

Cited by 32 publications

References 129 publications

Intraspecies Regulation of Ribonucleolytic Activity

Intraspecies Regulation of Ribonucleolytic Activity

The Relationship Between Relative Solvent Accessibility and Evolutionary Rate in Protein Evolution

Back to the future: Ribonuclease A

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