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Protein Engineering Design & Selection最新文献

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Engineering enzyme activity using an expanded amino acid alphabet. 利用扩展的氨基酸字母表工程酶活性。
IF 2.4 4区 生物学
Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzac013
Zachary Birch-Price, Christopher J Taylor, Mary Ortmayer, Anthony P Green
{"title":"Engineering enzyme activity using an expanded amino acid alphabet.","authors":"Zachary Birch-Price,&nbsp;Christopher J Taylor,&nbsp;Mary Ortmayer,&nbsp;Anthony P Green","doi":"10.1093/protein/gzac013","DOIUrl":"https://doi.org/10.1093/protein/gzac013","url":null,"abstract":"<p><p>Enzyme design and engineering strategies are typically constrained by the limited size of nature's genetic alphabet, comprised of only 20 canonical amino acids. In recent years, site-selective incorporation of non-canonical amino acids (ncAAs) via an expanded genetic code has emerged as a powerful means of inserting new functional components into proteins, with hundreds of structurally diverse ncAAs now available. Here, we highlight how the emergence of an expanded repertoire of amino acids has opened new avenues in enzyme design and engineering. ncAAs have been used to probe complex biological mechanisms, augment enzyme function and, most ambitiously, embed new catalytic mechanisms into protein active sites that would be challenging to access within the constraints of nature's genetic code. We predict that the studies reviewed in this article, along with further advances in genetic code expansion technology, will establish ncAA incorporation as an increasingly important tool for biocatalysis in the coming years.</p>","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2023-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9863031/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10576929","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 3
abYpap: improvements to the prediction of antibody VH/VL packing using gradient boosted regression. abYpap:使用梯度增强回归预测抗体V H/V L包装的改进。
IF 2.4 4区 生物学
Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzad021
Veronica A Boron, Andrew C R Martin
{"title":"abYpap: improvements to the prediction of antibody VH/VL packing using gradient boosted regression.","authors":"Veronica A Boron, Andrew C R Martin","doi":"10.1093/protein/gzad021","DOIUrl":"10.1093/protein/gzad021","url":null,"abstract":"<p><p>The Fv region of the antibody (comprising VH and VL domains) is the area responsible for target binding and thus the antibody's specificity. The orientation, or packing, of these two domains relative to each other influences the topography of the Fv region, and therefore can influence the antibody's binding affinity. We present abYpap, an improved method for predicting the packing angle between the VH and VL domains. With the large data set now available, we were able to expand greatly the number of features that could be used compared with our previous work. The machine-learning model was tuned for improved performance using 37 selected residues (previously 13) and also by including the lengths of the most variable 'complementarity determining regions' (CDR-L1, CDR-L2 and CDR-H3). Our method shows large improvements from the previous version, and also against other modeling approaches, when predicting the packing angle.</p>","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2023-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10719492/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"138453069","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Engineering cellulases for conversion of lignocellulosic biomass. 转化木质纤维素生物质的纤维素酶工程。
IF 2.4 4区 生物学
Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzad002
Yogesh B Chaudhari, Anikó Várnai, Morten Sørlie, Svein J Horn, Vincent G H Eijsink
{"title":"Engineering cellulases for conversion of lignocellulosic biomass.","authors":"Yogesh B Chaudhari, Anikó Várnai, Morten Sørlie, Svein J Horn, Vincent G H Eijsink","doi":"10.1093/protein/gzad002","DOIUrl":"10.1093/protein/gzad002","url":null,"abstract":"<p><p>Lignocellulosic biomass is a renewable source of energy, chemicals and materials. Many applications of this resource require the depolymerization of one or more of its polymeric constituents. Efficient enzymatic depolymerization of cellulose to glucose by cellulases and accessory enzymes such as lytic polysaccharide monooxygenases is a prerequisite for economically viable exploitation of this biomass. Microbes produce a remarkably diverse range of cellulases, which consist of glycoside hydrolase (GH) catalytic domains and, although not in all cases, substrate-binding carbohydrate-binding modules (CBMs). As enzymes are a considerable cost factor, there is great interest in finding or engineering improved and robust cellulases, with higher activity and stability, easy expression, and minimal product inhibition. This review addresses relevant engineering targets for cellulases, discusses a few notable cellulase engineering studies of the past decades and provides an overview of recent work in the field.</p>","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2023-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10394125/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9929667","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Engineering Candida boidinii formate dehydrogenase for activity with the non-canonical cofactor 3'-NADP(H). 利用非经典辅因子3'-NADP（H）对博伊迪尼假丝酵母甲酸脱氢酶进行活性改造。
IF 2.4 4区 生物学
Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzad009
Salomon Vainstein, Scott Banta
{"title":"Engineering Candida boidinii formate dehydrogenase for activity with the non-canonical cofactor 3'-NADP(H).","authors":"Salomon Vainstein,&nbsp;Scott Banta","doi":"10.1093/protein/gzad009","DOIUrl":"10.1093/protein/gzad009","url":null,"abstract":"<p><p>Oxidoreductases catalyze essential redox reactions, and many require a diffusible cofactor for electron transport, such as NAD(H). Non-canonical cofactor analogs have been explored as a means to create enzymatic reactions that operate orthogonally to existing metabolism. Here, we aimed to engineer the formate dehydrogenase from Candid boidinii (CbFDH) for activity with the non-canonical cofactor nicotinamide adenine dinucleotide 3'-phosphate (3'-NADP(H)). We used PyRosetta, the Cofactor Specificity Reversal Structural Analysis and Library Design (CSR-SALAD), and structure-guided saturation mutagenesis to identify mutations that enable CbFDH to use 3'-NADP+. Two single mutants, D195A and D195G, had the highest activities with 3'-NADP+, while the double mutant D195G/Y196S exhibited the highest cofactor selectivity reversal behavior. Steady state kinetic analyses were performed; the D195A mutant exhibited the highest KTS value with 3'-NADP+. This work compares the utility of computational approaches for cofactor specificity engineering while demonstrating the engineering of an important enzyme for novel non-canonical cofactor selectivity.</p>","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2023-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10146296","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Masked inverse folding with sequence transfer for protein representation learning. 用于蛋白质表示学习的带序列转移的掩模反向折叠。
IF 2.4 4区 生物学
Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzad015
Kevin K Yang, Niccolò Zanichelli, Hugh Yeh
{"title":"Masked inverse folding with sequence transfer for protein representation learning.","authors":"Kevin K Yang, Niccolò Zanichelli, Hugh Yeh","doi":"10.1093/protein/gzad015","DOIUrl":"10.1093/protein/gzad015","url":null,"abstract":"<p><p>Self-supervised pretraining on protein sequences has led to state-of-the art performance on protein function and fitness prediction. However, sequence-only methods ignore the rich information contained in experimental and predicted protein structures. Meanwhile, inverse folding methods reconstruct a protein's amino-acid sequence given its structure, but do not take advantage of sequences that do not have known structures. In this study, we train a masked inverse folding protein masked language model parameterized as a structured graph neural network. During pretraining, this model learns to reconstruct corrupted sequences conditioned on the backbone structure. We then show that using the outputs from a pretrained sequence-only protein masked language model as input to the inverse folding model further improves pretraining perplexity. We evaluate both of these models on downstream protein engineering tasks and analyze the effect of using information from experimental or predicted structures on performance.</p>","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2023-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"54232290","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Engineering of a phosphotriesterase with improved stability and enhanced activity for detoxification of the pesticide metabolite malaoxon. 一种具有改进的稳定性和增强的农药代谢产物马拉氧酮解毒活性的磷酸三酯酶的工程。
IF 2.4 4区 生物学
Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzad020
Laura Job, Anja Köhler, Mauricio Testanera, Benjamin Escher, Franz Worek, Arne Skerra
{"title":"Engineering of a phosphotriesterase with improved stability and enhanced activity for detoxification of the pesticide metabolite malaoxon.","authors":"Laura Job, Anja Köhler, Mauricio Testanera, Benjamin Escher, Franz Worek, Arne Skerra","doi":"10.1093/protein/gzad020","DOIUrl":"10.1093/protein/gzad020","url":null,"abstract":"<p><p>Organophosphorus (OP) pesticides are still widely applied but pose a severe toxicological threat if misused. For in vivo detoxification, the application of hydrolytic enzymes potentially offers a promising treatment. A well-studied example is the phosphotriesterase of Brevundimonas diminuta (BdPTE). Whereas wild-type BdPTE can hydrolyse pesticides like paraoxon, chlorpyrifos-oxon and mevinphos with high catalytic efficiencies, kcat/KM >2 × 107 M-1 min-1, degradation of malaoxon is unsatisfactory (kcat/KM ≈ 1 × 104 M-1 min-1). Here, we report the rational engineering of BdPTE mutants with improved properties and their efficient production in Escherichia coli. As result, the mutant BdPTE(VRNVVLARY) exhibits 37-fold faster malaoxon hydrolysis (kcat/KM = 4.6 × 105 M-1 min-1), together with enhanced expression yield, improved thermal stability and reduced susceptibility to oxidation. Therefore, this BdPTE mutant constitutes a powerful candidate to develop a biocatalytic antidote for the detoxification of this common pesticide metabolite as well as related OP compounds.</p>","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2023-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"71523393","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Correction to: The variable conversion of neutralizing anti-SARS-CoV-2 single-chain antibodies to IgG provides insight into RBD epitope accessibility. 更正：中和抗严重急性呼吸系统综合征冠状病毒2型单链抗体向IgG的可变转化提供了对RBD表位可及性的深入了解。
IF 2.4 4区 生物学
Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzad018
{"title":"Correction to: The variable conversion of neutralizing anti-SARS-CoV-2 single-chain antibodies to IgG provides insight into RBD epitope accessibility.","authors":"","doi":"10.1093/protein/gzad018","DOIUrl":"10.1093/protein/gzad018","url":null,"abstract":"","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2023-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10628328/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"71488998","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
A protein engineering approach toward understanding FKBP51 conformational dynamics and mechanisms of ligand binding. 一种了解FKBP51构象动力学和配体结合机制的蛋白质工程方法。
IF 2.4 4区 生物学
Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzad014
Jorge A Lerma Romero, Christian Meyners, Nicole Rupp, Felix Hausch, Harald Kolmar
{"title":"A protein engineering approach toward understanding FKBP51 conformational dynamics and mechanisms of ligand binding.","authors":"Jorge A Lerma Romero,&nbsp;Christian Meyners,&nbsp;Nicole Rupp,&nbsp;Felix Hausch,&nbsp;Harald Kolmar","doi":"10.1093/protein/gzad014","DOIUrl":"10.1093/protein/gzad014","url":null,"abstract":"<p><p>Most proteins are flexible molecules that coexist in an ensemble of several conformations. Point mutations in the amino acid sequence of a protein can trigger structural changes that drive the protein population to a conformation distinct from the native state. Here, we report a protein engineering approach to better understand protein dynamics and ligand binding of the FK506-binding protein 51 (FKBP51), a prospective target for stress-related diseases, metabolic disorders, some types of cancers and chronic pain. By randomizing selected regions of its ligand-binding domain and sorting yeast display libraries expressing these variants, mutants with high affinity to conformation-specific FKBP51 selective ligands were identified. These improved mutants are valuable tools for the discovery of novel selective ligands that preferentially and specifically bind the FKBP51 active site in its open conformation state. Moreover, they will help us understand the conformational dynamics and ligand binding mechanics of the FKBP51 binding pocket.</p>","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2023-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"71415303","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
Non-canonical amino acids as a tool for the thermal stabilization of enzymes. 作为酶热稳定工具的非典型氨基酸。
IF 2.6 4区 生物学
Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzad003
Tim Lugtenburg, Alejandro Gran-Scheuch, Ivana Drienovská
{"title":"Non-canonical amino acids as a tool for the thermal stabilization of enzymes.","authors":"Tim Lugtenburg, Alejandro Gran-Scheuch, Ivana Drienovská","doi":"10.1093/protein/gzad003","DOIUrl":"10.1093/protein/gzad003","url":null,"abstract":"<p><p>Biocatalysis has become a powerful alternative for green chemistry. Expanding the range of amino acids used in protein biosynthesis can improve industrially appealing properties such as enantioselectivity, activity and stability. This review will specifically delve into the thermal stability improvements that non-canonical amino acids (ncAAs) can confer to enzymes. Methods to achieve this end, such as the use of halogenated ncAAs, selective immobilization and rational design, will be discussed. Additionally, specific enzyme design considerations using ncAAs are discussed along with the benefits and limitations of the various approaches available to enhance the thermal stability of enzymes.</p>","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":null,"pages":null},"PeriodicalIF":2.6,"publicationDate":"2023-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10064326/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9347753","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
An easy-to-use high-throughput selection system for the discovery of recombinant protein binders from alternative scaffold libraries. 一种易于使用的高通量选择系统，用于从替代支架库中发现重组蛋白结合物。
IF 2.4 4区 生物学
Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzad011
Marit Möller, Malin Jönsson, Magnus Lundqvist, Blenda Hedin, Louise Larsson, Emma Larsson, Johan Rockberg, Mathias Uhlén, Sarah Lindbo, Hanna Tegel, Sophia Hober
{"title":"An easy-to-use high-throughput selection system for the discovery of recombinant protein binders from alternative scaffold libraries.","authors":"Marit Möller,&nbsp;Malin Jönsson,&nbsp;Magnus Lundqvist,&nbsp;Blenda Hedin,&nbsp;Louise Larsson,&nbsp;Emma Larsson,&nbsp;Johan Rockberg,&nbsp;Mathias Uhlén,&nbsp;Sarah Lindbo,&nbsp;Hanna Tegel,&nbsp;Sophia Hober","doi":"10.1093/protein/gzad011","DOIUrl":"10.1093/protein/gzad011","url":null,"abstract":"<p><p>Selection by phage display is a popular and widely used technique for the discovery of recombinant protein binders from large protein libraries for therapeutic use. The protein library is displayed on the surface of bacteriophages which are amplified using bacteria, preferably Escherichia coli, to enrich binders in several selection rounds. Traditionally, the so-called panning procedure during which the phages are incubated with the target protein, washed and eluted is done manually, limiting the throughput. High-throughput systems with automated panning already in use often require high-priced equipment. Moreover, the bottleneck of the selection process is usually the screening and characterization. Therefore, having a high-throughput panning procedure without a scaled screening platform does not necessarily increase the discovery rate. Here, we present an easy-to-use high-throughput selection system with automated panning using cost-efficient equipment integrated into a workflow with high-throughput sequencing and a tailored screening step using biolayer-interferometry. The workflow has been developed for selections using two recombinant libraries, ADAPT (Albumin-binding domain-derived affinity proteins) and CaRA (Calcium-regulated affinity) and has been evaluated for three new targets. The newly established semi-automated system drastically reduced the hands-on time and increased robustness while the selection outcome, when compared to manual handling, was very similar in deep sequencing analysis and generated binders in the nanomolar affinity range. The developed selection system has shown to be highly versatile and has the potential to be applied to other binding domains for the discovery of new protein binders.</p>","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":null,"pages":null},"PeriodicalIF":2.4,"publicationDate":"2023-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/7f/95/gzad011.PMC10545973.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10278675","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
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