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Jorge Fernandez-de-Cossio-Diaz Guido Uguzzoni Andrea Pagnani 《Molecular biology and evolution》2021,38(1):318
The recent technological advances underlying the screening of large combinatorial libraries in high-throughput mutational scans deepen our understanding of adaptive protein evolution and boost its applications in protein design. Nevertheless, the large number of possible genotypes requires suitable computational methods for data analysis, the prediction of mutational effects, and the generation of optimized sequences. We describe a computational method that, trained on sequencing samples from multiple rounds of a screening experiment, provides a model of the genotype–fitness relationship. We tested the method on five large-scale mutational scans, yielding accurate predictions of the mutational effects on fitness. The inferred fitness landscape is robust to experimental and sampling noise and exhibits high generalization power in terms of broader sequence space exploration and higher fitness variant predictions. We investigate the role of epistasis and show that the inferred model provides structural information about the 3D contacts in the molecular fold. 相似文献
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Susanne A. Kraemer Katharina B. Böndel Robert W. Ness Peter D. Keightley Nick Colegrave 《Evolution; international journal of organic evolution》2017,71(12):2918-2929
Although all genetic variation ultimately stems from mutations, their properties are difficult to study directly. Here, we used multiple mutation accumulation (MA) lines derived from five genetic backgrounds of the green algae Chlamydomonas reinhardtii that have been previously subjected to whole genome sequencing to investigate the relationship between the number of spontaneous mutations and change in fitness from a nonevolved ancestor. MA lines were on average less fit than their ancestors and we detected a significantly negative correlation between the change in fitness and the total number of accumulated mutations in the genome. Likewise, the number of mutations located within coding regions significantly and negatively impacted MA line fitness. We used the fitness data to parameterize a maximum likelihood model to estimate discrete categories of mutational effects, and found that models containing one to two mutational effect categories (one neutral and one deleterious category) fitted the data best. However, the best‐fitting mutational effects models were highly dependent on the genetic background of the ancestral strain. 相似文献
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Xiao‐Lin Chu Da‐Yong Zhang Angus Buckling Quan‐Guo Zhang 《Journal of evolutionary biology》2020,33(8):1020-1027
Temperature determines the rates of all biochemical and biophysical processes, and is also believed to be a key driver of macroevolutionary patterns. It is suggested that physiological constraints at low temperatures may diminish the fitness advantages of otherwise beneficial mutations; by contrast, relatively high, benign, temperatures allow beneficial mutations to efficiently show their phenotypic effects. To experimentally test this “mutational effects” mechanism, we examined the fitness effects of mutations across a temperature gradient using bacterial genotypes from the early stage of a mutation accumulation experiment with Escherichia coli. While the incidence of beneficial mutations did not significantly change across environmental temperatures, the number of mutations that conferred strong beneficial fitness effects was greater at higher temperatures. The results therefore support the hypothesis that warmer temperatures increase the chance and magnitude of positive selection, with implications for explaining the geographic patterns in evolutionary rates and understanding contemporary evolution under global warming. 相似文献
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Amino acids fulfil a diverse range of roles in proteins, each utilising its chemical properties in different ways in different contexts to create required functions. For example, cysteines form disulphide or hydrogen bonds in different circumstances and charged amino acids do not always make use of their charge. The repertoire of amino acid functions and the frequency at which they occur in proteins remains understudied. Measuring large numbers of mutational consequences, which can elucidate the role an amino acid plays, was prohibitively time‐consuming until recent developments in deep mutational scanning. In this study, we gathered data from 28 deep mutational scanning studies, covering 6,291 positions in 30 proteins, and used the consequences of mutation at each position to define a mutational landscape. We demonstrated rich relationships between this landscape and biophysical or evolutionary properties. Finally, we identified 100 functional amino acid subtypes with a data‐driven clustering analysis and studied their features, including their frequencies and chemical properties such as tolerating polarity, hydrophobicity or being intolerant of charge or specific amino acids. The mutational landscape and amino acid subtypes provide a foundational catalogue of amino acid functional diversity, which will be refined as the number of studied protein positions increases. 相似文献
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Fitness effects of mutations may generally depend on temperature that influences all rate-limiting biophysical and biochemical processes. Earlier studies suggested that high temperatures may increase the availability of beneficial mutations (‘more beneficial mutations’), or allow beneficial mutations to show stronger fitness effects (‘stronger beneficial mutation effects’). The ‘more beneficial mutations’ scenario would inevitably be associated with increased proportion of conditionally beneficial mutations at higher temperatures. This in turn predicts that populations in warm environments show faster evolutionary adaptation but suffer fitness loss when faced with cold conditions, and those evolving in cold environments become thermal-niche generalists (‘hotter is narrower’). Under the ‘stronger beneficial mutation effects’ scenario, populations evolving in warm environments would show faster adaptation without fitness costs in cold environments, leading to a ‘hotter is (universally) better’ pattern in thermal niche adaptation. We tested predictions of the two competing hypotheses using an experimental evolution study in which populations of two model bacterial species, Escherichia coli and Pseudomonas fluorescens, evolved for 2400 generations at three experimental temperatures. Results of reciprocal transplant experiments with our P. fluorescens populations were largely consistent with the ‘hotter is narrower’ prediction. Results from the E. coli populations clearly suggested stronger beneficial mutation effects at higher assay temperatures, but failed to detect faster adaptation in populations evolving in warmer experimental environments (presumably because of limitation in the supply of genetic variation). Our results suggest that the influence of temperature on mutational effects may provide insight into the patterns of thermal niche adaptation and population diversification across thermal conditions. 相似文献
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作为细胞结构与功能的中心参与者,蛋白质一直是生命科学研究的中心主题。分析蛋白质序列变异对其结构、功能的影响,是研究蛋白的重要手段之一。近年一种称为深度突变扫描(deep mutational scanning,DMS)的技术被广泛应用于蛋白研究领域,其通过高丰度DNA文库在蛋白特定区域平行引入成千上万种突变,经筛选后,利用高通量测序为每一种突变打分,从而揭示序列与功能之间的相关性。深度突变扫描以其高通量、快速简易、节省人工等特点,已经成为蛋白质功能研究以及蛋白工程改造的一种重要方法,目前已在蛋白进化、抗体改造、致病突变鉴定等蛋白研究的多个领域广泛应用。本综述简要概括了深度突变扫描技术的原理,重点介绍了其在哺乳动物细胞中的应用,同时分析了目前的技术瓶颈,旨在为相关研究提供参考。 相似文献
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Allison J. Greaney Tyler N. Starr Pavlo Gilchuk Seth J. Zost Elad Binshtein Andrea N. Loes Sarah K. Hilton John Huddleston Rachel Eguia Katharine H.D. Crawford Adam S. Dingens Rachel S. Nargi Rachel E. Sutton Naveenchandra Suryadevara Paul W. Rothlauf Zhuoming Liu Sean P.J. Whelan Robert H. Carnahan Jesse D. Bloom 《Cell host & microbe》2021,29(1):44-57.e9
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Elena SF Wilke CO Ofria C Lenski RE 《Evolution; international journal of organic evolution》2007,61(3):666-674
It is often assumed that the efficiency of selection for mutational robustness would be proportional to mutation rate and population size, thus being inefficient in small populations. However, Krakauer and Plotkin (2002) hypothesized that selection in small populations would favor robustness mechanisms, such as redundancy, that mask the effect of deleterious mutations. In large populations, by contrast, selection is more effective at removing deleterious mutants and fitness would be improved by eliminating mechanisms that mask the effect of deleterious mutations and thus impede their removal. Here, we test whether these predictions are supported in experiments with evolving populations of digital organisms. Digital organisms are self-replicating programs that inhabit a virtual world inside a computer. Like their organic counterparts, digital organisms mutate, compete, evolve, and adapt by natural selection to their environment. In this study, 160 populations evolved at different combinations of mutation rate and population size. After 10(4) generations, we measured the mutational robustness of the most abundant genotype in each population. Mutational robustness tended to increase with mutation rate and to decline with population size, although the dependence with population size was in part mediated by a negative relationship between fitness and robustness. These results are independent of whether genomes were constrained to their original length or allowed to change in size. 相似文献
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《Journal of molecular biology》2023,435(18):168209
Characterizing the effects of mutations on stability is critical for understanding the function and evolution of proteins and improving their biophysical properties. High throughput folding and abundance assays have been successfully used to characterize missense mutations associated with reduced stability. However, screening for increased thermodynamic stability is more challenging since such mutations are rarer and their impact on assay readout is more subtle. Here, a multiplex assay for high throughput screening of protein folding was developed by combining deep mutational scanning, fluorescence-activated cell sorting, and deep sequencing. By analyzing a library of 2000 variants of Adenylate kinase we demonstrate that the readout of the method correlates with stability and that mutants with up to 13 °C increase in thermal melting temperature could be identified with low false positive rate. The discovery of many stabilizing mutations also enabled the analysis of general substitution patterns associated with increased stability in Adenylate kinase. This high throughput method to identify stabilizing mutations can be combined with functional screens to identify mutations that improve both stability and activity. 相似文献
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Parul Johri Jean-Francois Gout Thomas G Doak Michael Lynch 《Molecular biology and evolution》2022,39(6)
Whole-genome duplications (WGDs) have occurred in many eukaryotic lineages. However, the underlying evolutionary forces and molecular mechanisms responsible for the long-term retention of gene duplicates created by WGDs are not well understood. We employ a population-genomic approach to understand the selective forces acting on paralogs and investigate ongoing duplicate-gene loss in multiple species of Paramecium that share an ancient WGD. We show that mutations that abolish protein function are more likely to be segregating in retained WGD paralogs than in single-copy genes, most likely because of ongoing nonfunctionalization post-WGD. This relaxation of purifying selection occurs in only one WGD paralog, accompanied by the gradual fixation of nonsynonymous mutations and reduction in levels of expression, and occurs over a long period of evolutionary time, “marking” one locus for future loss. Concordantly, the fitness effects of new nonsynonymous mutations and frameshift-causing indels are significantly more deleterious in the highly expressed copy compared with their paralogs with lower expression. Our results provide a novel mechanistic model of gene duplicate loss following WGDs, wherein selection acts on the sum of functional activity of both duplicate genes, allowing the two to wander in expression and functional space, until one duplicate locus eventually degenerates enough in functional efficiency or expression that its contribution to total activity is too insignificant to be retained by purifying selection. Retention of duplicates by such mechanisms predicts long times to duplicate-gene loss, which should not be falsely attributed to retention due to gain/change in function. 相似文献
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Andrew M. Wollacott Luke N. Robinson Boopathy Ramakrishnan Hamid Tissire Karthik Viswanathan Zachary Shriver Gregory J. Babcock 《Journal of molecular recognition : JMR》2019,32(7)
IgA nephropathy (IgAN) is the most prevalent cause of primary glomerular disease worldwide, and the cytokine A PRoliferation‐Inducing Ligand (APRIL) is emerging as a key player in IgAN pathogenesis and disease progression. For a panel of anti‐human APRIL antibodies with known antagonistic activity, we sought to define their structural mode of engagement to understand molecular mechanisms of action and aid rational antibody engineering. Reliable computational prediction of antibody‐antigen complexes remains challenging, and experimental methods such as X‐ray co‐crystallography and cryoEM have considerable technical, resource, and throughput barriers. To overcome these limitations, we implemented an integrated and accessible experimental‐computational workflow to more accurately predict structures of antibody‐APRIL complexes. Specifically, a yeast surface display library encoding site‐saturation mutagenized surface positions of APRIL was screened against a panel of anti‐APRIL antibodies to rapidly obtain a comprehensive biochemical profile of mutational impact on binding for each antibody. The experimentally derived mutational profile data were used as quantitative constraints in a computational docking workflow optimized for antibodies, resulting in robust structural models of antibody‐antigen complexes. The model results were confirmed by solving the cocrystal structure of one antibody‐APRIL complex, which revealed strong agreement with our model. The models were used to rationally select and engineer one antibody for cross‐species APRIL binding, which quite often aids further testing in relevant animal models. Collectively, we demonstrate a rapid, integrated computational‐experimental approach to robustly predict antibody‐antigen structures information, which can aid rational antibody engineering and provide insights into molecular mechanisms of action. 相似文献
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Jack Da Silva 《Evolution; international journal of organic evolution》2010,64(4):1160-1165
The mutational landscape model of adaptive sequence evolution has been used to explain an unexpected strong positive linear relationship between marginal fitness and mean site‐specific amino acid frequency in the functionally important HIV‐1 gp120 V3 protein region. The model predicts a positive linear relationship between the probability that a particular beneficial allele, among several, is the next to spread to fixation during an adaptive walk, its transition probability, and the allele's selection coefficient. Here, stochastic simulation is used to confirm the intuition that the linear relationship between transition probabilities and selection coefficients, predicted by the model, should, under fluctuating selection, produce a linear relationship between allele frequency, averaged across populations, and fitness. In addition, these relationships hold for the effective population size and mutation rate of HIV‐1 and for the moderately strong selection observed for V3. A survey of the strength of mutation for diverse organisms suggests that these relationships may be widely applicable. 相似文献
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Yusuf Talha Tamer Ilona Gaszek Marinelle Rodrigues Fatma Sevde Coskun Michael Farid Andrew Y Koh William Russ Erdal Toprak 《Molecular biology and evolution》2021,38(10):4493
Bacteriophages and bacterial toxins are promising antibacterial agents to treat infections caused by multidrug-resistant (MDR) bacteria. In fact, bacteriophages have recently been successfully used to treat life-threatening infections caused by MDR bacteria (Schooley RT, Biswas B, Gill JJ, Hernandez-Morales A, Lancaster J, Lessor L, Barr JJ, Reed SL, Rohwer F, Benler S, et al. 2017. Development and use of personalized bacteriophage-based therapeutic cocktails to treat a patient with a disseminated resistant Acinetobacter baumannii infection. Antimicrob Agents Chemother. 61(10); Chan BK, Turner PE, Kim S, Mojibian HR, Elefteriades JA, Narayan D. 2018. Phage treatment of an aortic graft infected with Pseudomonas aeruginosa. Evol Med Public Health. 2018(1):60–66; Petrovic Fabijan A, Lin RCY, Ho J, Maddocks S, Ben Zakour NL, Iredell JR, Westmead Bacteriophage Therapy Team. 2020. Safety of bacteriophage therapy in severe Staphylococcus aureus infection. Nat Microbiol. 5(3):465–472). One potential problem with using these antibacterial agents is the evolution of resistance against them in the long term. Here, we studied the fitness landscape of the Escherichia coli TolC protein, an outer membrane efflux protein that is exploited by a pore forming toxin called colicin E1 and by TLS phage (Pagie L, Hogeweg P. 1999. Colicin diversity: a result of eco-evolutionary dynamics. J Theor Biol. 196(2):251–261; Andersen C, Hughes C, Koronakis V. 2000. Chunnel vision. Export and efflux through bacterial channel-tunnels. EMBO Rep. 1(4):313–318; Koronakis V, Andersen C, Hughes C. 2001. Channel-tunnels. Curr Opin Struct Biol. 11(4):403–407; Czaran TL, Hoekstra RF, Pagie L. 2002. Chemical warfare between microbes promotes biodiversity. Proc Natl Acad Sci U S A. 99(2):786–790; Cascales E, Buchanan SK, Duché D, Kleanthous C, Lloubès R, Postle K, Riley M, Slatin S, Cavard D. 2007. Colicin biology. Microbiol Mol Biol Rev. 71(1):158–229). By systematically assessing the distribution of fitness effects of ∼9,000 single amino acid replacements in TolC using either positive (antibiotics and bile salts) or negative (colicin E1 and TLS phage) selection pressures, we quantified evolvability of the TolC. We demonstrated that the TolC is highly optimized for the efflux of antibiotics and bile salts. In contrast, under colicin E1 and TLS phage selection, TolC sequence is very sensitive to mutations. Finally, we have identified a large set of mutations in TolC that increase resistance of E. coli against colicin E1 or TLS phage without changing antibiotic susceptibility of bacterial cells. Our findings suggest that TolC is a highly evolvable target under negative selection which may limit the potential clinical use of bacteriophages and bacterial toxins if evolutionary aspects are not taken into account. 相似文献
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Atina G Cote Jennifer Knapp Marta Verby Joseph C Mellor Yingzhou Wu Carles Pons Cassandra Wong Natascha van Lieshout Fan Yang Murat Tasan Guihong Tan Shan Yang Douglas M Fowler Robert Nussbaum Jesse D Bloom Marc Vidal David E Hill Patrick Aloy Frederick P Roth 《Molecular systems biology》2017,13(12)
Although we now routinely sequence human genomes, we can confidently identify only a fraction of the sequence variants that have a functional impact. Here, we developed a deep mutational scanning framework that produces exhaustive maps for human missense variants by combining random codon mutagenesis and multiplexed functional variation assays with computational imputation and refinement. We applied this framework to four proteins corresponding to six human genes: UBE2I (encoding SUMO E2 conjugase), SUMO1 (small ubiquitin‐like modifier), TPK1 (thiamin pyrophosphokinase), and CALM1/2/3 (three genes encoding the protein calmodulin). The resulting maps recapitulate known protein features and confidently identify pathogenic variation. Assays potentially amenable to deep mutational scanning are already available for 57% of human disease genes, suggesting that DMS could ultimately map functional variation for all human disease genes. 相似文献
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Clifford Zeyl Melissa Mizesko J. Arjan G. M. de Visser 《Evolution; international journal of organic evolution》2001,55(5):909-917
Abstract.— In small or repeatedly bottlenecked populations, mutations are expected to accumulate by genetic drift, causing fitness declines. In mutational meltdown models, such fitness declines further reduce population size, thus accelerating additional mutation accumulation and leading to extinction. Because the rate of mutation accumulation is determined partly by the mutation rate, the risk and rate of meltdown are predicted to increase with increasing mutation rate. We established 12 replicate populations of Saccharomyces cerevisiae from each of two isogenic strains whose genomewide mutation rates differ by approximately two orders of magnitude. Each population was transferred daily by a fixed dilution that resulted in an effective population size near 250. Fitness declines that reduce growth rates were expected to reduce the numbers of cells transferred after dilution, thus reducing population size and leading to mutational meltdown. Through 175 daily transfers and approximately 2900 generations, two extinctions occurred, both in populations with elevated mutation rates. For one of these populations there is direct evidence that extinction resulted from mutational meltdown: Extinction immediately followed a major fitness decline, and it recurred consistently in replicate populations reestablished from a sample frozen after this fitness decline, but not in populations founded from a predecline sample. Wild‐type populations showed no trend to decrease in size and, on average, they increased in fitness. 相似文献
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Julia Zischewski Lucia Perez Ludovic Bassié Riad Nadi Giobbe Forni Sarah Boyd Lade Erika Soto Xin Jin Vicente Medina Gemma Villorbina Pilar Muñoz Gemma Farré Rainer Fischer Richard M. Twyman Teresa Capell Paul Christou Stefan Schillberg 《Plant biotechnology journal》2016,14(12):2203-2216
The CRISPR/Cas9 system and related RNA‐guided endonucleases can introduce double‐strand breaks (DSBs) at specific sites in the genome, allowing the generation of targeted mutations in one or more genes as well as more complex genomic rearrangements. Modifications of the canonical CRISPR/Cas9 system from Streptococcus pyogenes and the introduction of related systems from other bacteria have increased the diversity of genomic sites that can be targeted, providing greater control over the resolution of DSBs, the targeting efficiency (frequency of on‐target mutations), the targeting accuracy (likelihood of off‐target mutations) and the type of mutations that are induced. Although much is now known about the principles of CRISPR/Cas9 genome editing, the likelihood of different outcomes is species‐dependent and there have been few comparative studies looking at the basis of such diversity. Here we critically analyse the activity of CRISPR/Cas9 and related systems in different plant species and compare the outcomes in animals and microbes to draw broad conclusions about the design principles required for effective genome editing in different organisms. These principles will be important for the commercial development of crops, farm animals, animal disease models and novel microbial strains using CRISPR/Cas9 and other genome‐editing tools. 相似文献