Protein Engineering Design & Selection最新文献_第4页

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

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, Malin Jönsson, Magnus Lundqvist, Blenda Hedin, Louise Larsson, Emma Larsson, Johan Rockberg, Mathias Uhlén, Sarah Lindbo, Hanna Tegel, Sophia Hober","doi":"10.1093/protein/gzad011","DOIUrl":"10.1093/protein/gzad011","url":null,"abstract":"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.","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":" ","pages":""},"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

Data-driven enzyme engineering to identify function-enhancing enzymes. 数据驱动的酶工程，以识别功能增强酶。

IF 2.4 4区生物学

Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzac009

Yaoyukun Jiang, Xinchun Ran, Zhongyue J Yang

{"title":"Data-driven enzyme engineering to identify function-enhancing enzymes.","authors":"Yaoyukun Jiang, Xinchun Ran, Zhongyue J Yang","doi":"10.1093/protein/gzac009","DOIUrl":"10.1093/protein/gzac009","url":null,"abstract":"Identifying function-enhancing enzyme variants is a 'holy grail' challenge in protein science because it will allow researchers to expand the biocatalytic toolbox for late-stage functionalization of drug-like molecules, environmental degradation of plastics and other pollutants, and medical treatment of food allergies. Data-driven strategies, including statistical modeling, machine learning, and deep learning, have largely advanced the understanding of the sequence-structure-function relationships for enzymes. They have also enhanced the capability of predicting and designing new enzymes and enzyme variants for catalyzing the transformation of new-to-nature reactions. Here, we reviewed the recent progresses of data-driven models that were applied in identifying efficiency-enhancing mutants for catalytic reactions. We also discussed existing challenges and obstacles faced by the community. Although the review is by no means comprehensive, we hope that the discussion can inform the readers about the state-of-the-art in data-driven enzyme engineering, inspiring more joint experimental-computational efforts to develop and apply data-driven modeling to innovate biocatalysts for synthetic and pharmaceutical applications.","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":"36 ","pages":""},"PeriodicalIF":2.4,"publicationDate":"2023-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10365845/pdf/gzac009.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10316967","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

The variable conversion of neutralizing anti-SARS-CoV-2 single-chain antibodies to IgG provides insight into RBD epitope accessibility. 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 2.6 4区生物学

Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzad008

Matthew R Chang, Hanzhong Ke, Laura Losada Miguéns, Christian Coherd, Katrina Nguyen, Maliwan Kamkaew, Rebecca Johnson, Nadia Storm, Anna Honko, Quan Zhu, Anthony Griffiths, Wayne A Marasco

{"title":"The variable conversion of neutralizing anti-SARS-CoV-2 single-chain antibodies to IgG provides insight into RBD epitope accessibility.","authors":"Matthew R Chang, Hanzhong Ke, Laura Losada Miguéns, Christian Coherd, Katrina Nguyen, Maliwan Kamkaew, Rebecca Johnson, Nadia Storm, Anna Honko, Quan Zhu, Anthony Griffiths, Wayne A Marasco","doi":"10.1093/protein/gzad008","DOIUrl":"10.1093/protein/gzad008","url":null,"abstract":"Monoclonal antibody (mAb) therapies have rapidly become a powerful class of therapeutics with applications covering a diverse range of clinical indications. Though most widely used for the treatment of cancer, mAbs are also playing an increasing role in the defense of viral infections, most recently with palivizumab for prevention and treatment of severe RSV infections in neonatal and pediatric populations. In addition, during the COVID-19 pandemic, mAbs provided a bridge to the rollout of vaccines; however, their continued role as a therapeutic option for those at greatest risk of severe disease has become limited due to the emergence of neutralization resistant Omicron variants. Although there are many techniques for the identification of mAbs, including single B cell cloning and immunization of genetically engineered mice, the low cost, rapid throughput and technological simplicity of antibody phage display has led to its widespread adoption in mAb discovery efforts. Here we used our 27-billion-member naïve single-chain antibody (scFv) phage library to identify a panel of neutralizing anti-SARS-CoV-2 scFvs targeting diverse epitopes on the receptor binding domain (RBD). Although typically a routine process, we found that upon conversion to IgG, a number of our most potent clones failed to maintain their neutralization potency. Kinetic measurements confirmed similar affinity to the RBD; however, mechanistic studies provide evidence that the loss of neutralization is a result of structural limitations likely arising from initial choice of panning antigen. Thus this work highlights a risk of scFv-phage panning to mAb conversion and the importance of initial antigen selection.","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":"36 ","pages":""},"PeriodicalIF":2.6,"publicationDate":"2023-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10505556/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10355296","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

De novo design of a polycarbonate hydrolase. 聚碳酸酯水解酶的新设计。

IF 2.4 4区生物学

Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzad022

Laura H Holst, Niklas G Madsen, Freja T Toftgård, Freja Rønne, Ioana-Malina Moise, Evamaria I Petersen, Peter Fojan

引用次数: 0

Contributions from ClpS surface residues in modulating N-terminal peptide binding and their implications for NAAB development. ClpS表面残基在调节N-末端肽结合中的贡献及其对NAAB发育的影响。

IF 2.4 4区生物学

Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzad007

Nicholas Callahan, William B Siegall, Christina Bergonzo, John P Marino, Zvi Kelman

{"title":"Contributions from ClpS surface residues in modulating N-terminal peptide binding and their implications for NAAB development.","authors":"Nicholas Callahan, William B Siegall, Christina Bergonzo, John P Marino, Zvi Kelman","doi":"10.1093/protein/gzad007","DOIUrl":"10.1093/protein/gzad007","url":null,"abstract":"Numerous technologies are currently in development for use in next-generation protein sequencing platforms. A notable published approach employs fluorescently-tagged binding proteins to identity the N-terminus of immobilized peptides, in-between rounds of digestion. This approach makes use of N-terminal amino acid binder (NAAB) proteins, which would identify amino acids by chemical and shape complementarity. One source of NAABs is the ClpS protein family, which serve to recruit proteins to bacterial proteosomes based on the identity of the N-terminal amino acid. In this study, a Thermosynechococcus vestitus (also known as Thermosynechococcus elongatus) ClpS2 protein was used as the starting point for direct evolution of an NAAB with affinity and specificity for N-terminal leucine. Enriched variants were analyzed and shown to improve the interaction between the ClpS surface and the peptide chain, without increasing promiscuity. Interestingly, interactions were found that were unanticipated which favor different charged residues located at position 5 from the N-terminus of a target peptide.","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":"36 ","pages":""},"PeriodicalIF":2.4,"publicationDate":"2023-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10076735","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

Enzyme redesign and genetic code expansion. 酶的重新设计和遗传密码的扩展。

IF 2.4 4区生物学

Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzad017

Vaitea Opuu, Thomas Simonson

引用次数: 0

Analysis of conformational stability of interacting residues in protein binding interfaces. 蛋白质结合界面中相互作用残基构象稳定性的分析。

IF 2.4 4区生物学

Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzad016

Varun M Chauhan, Robert J Pantazes

引用次数: 0

Strategies for enriching and characterizing proteins with inhibitory properties on the yeast surface. 富集和鉴定酵母表面具有抑制特性的蛋白质的策略。

IF 2.4 4区生物学

Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzac017

Arlinda Rezhdo, Catherine T Lessard, Mariha Islam, James A Van Deventer

{"title":"Strategies for enriching and characterizing proteins with inhibitory properties on the yeast surface.","authors":"Arlinda Rezhdo, Catherine T Lessard, Mariha Islam, James A Van Deventer","doi":"10.1093/protein/gzac017","DOIUrl":"10.1093/protein/gzac017","url":null,"abstract":"Display technologies are powerful tools for discovering binding proteins against a broad range of biological targets. However, it remains challenging to adapt display technologies for the discovery of proteins that inhibit the enzymatic activities of targets. Here, we investigate approaches for discovering and characterizing inhibitory antibodies in yeast display format using a well-defined series of constructs and the target matrix metalloproteinase-9. Three previously reported antibodies were used to create model libraries consisting of inhibitory, non-inhibitory, and non-binding constructs. Conditions that preferentially enrich for inhibitory clones were identified for both magnetic bead-based enrichments and fluorescence-activated cell sorting. Half maximal inhibitory concentration (IC50) was obtained through yeast titration assays. The IC50 of the inhibitory antibody obtained in yeast display format falls within the confidence interval of the IC50 value determined in soluble form. Overall, this study identifies strategies for the discovery and characterization of inhibitory clones directly in yeast display format.","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":"36 ","pages":""},"PeriodicalIF":2.4,"publicationDate":"2023-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10365883/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10244236","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

Physics-based approach to extend a de novo TIM barrel with rationally designed helix-loop-helix motifs. 基于物理的方法，以合理设计的螺旋-环-螺旋图案扩展全新TIM桶。

IF 2.4 4区生物学

Protein Engineering Design & Selection Pub Date : 2023-01-21 DOI: 10.1093/protein/gzad012

Sina Kordes, Julian Beck, Sooruban Shanmugaratnam, Merle Flecks, Birte Höcker

{"title":"Physics-based approach to extend a de novo TIM barrel with rationally designed helix-loop-helix motifs.","authors":"Sina Kordes, Julian Beck, Sooruban Shanmugaratnam, Merle Flecks, Birte Höcker","doi":"10.1093/protein/gzad012","DOIUrl":"10.1093/protein/gzad012","url":null,"abstract":"Computational protein design promises the ability to build tailor-made proteins de novo. While a range of de novo proteins have been constructed so far, the majority of these designs have idealized topologies that lack larger cavities which are necessary for the incorporation of small molecule binding sites or enzymatic functions. One attractive target for enzyme design is the TIM-barrel fold, due to its ubiquity in nature and capability to host versatile functions. With the successful de novo design of a 4-fold symmetric TIM barrel, sTIM11, an idealized, minimalistic scaffold was created. In this work, we attempted to extend this de novo TIM barrel by incorporating a helix-loop-helix motif into its βα-loops by applying a physics-based modular design approach using Rosetta. Further diversification was performed by exploiting the symmetry of the scaffold to integrate two helix-loop-helix motifs into the scaffold. Analysis with AlphaFold2 and biochemical characterization demonstrate the formation of additional α-helical secondary structure elements supporting the successful extension as intended.","PeriodicalId":54543,"journal":{"name":"Protein Engineering Design & Selection","volume":" ","pages":""},"PeriodicalIF":2.4,"publicationDate":"2023-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10287313","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