D. Travis Gallagher, Natalia Oganesyan, Andrew Lees
{"title":"疫苗载体蛋白CRM197的单体晶体结构及其对疫苗开发的意义","authors":"D. Travis Gallagher, Natalia Oganesyan, Andrew Lees","doi":"10.1107/S2053230X23002364","DOIUrl":null,"url":null,"abstract":"<p>CRM<sub>197</sub> is a genetically detoxified mutant of diphtheria toxin (DT) that is widely used as a carrier protein in conjugate vaccines. Protective immune responses to several bacterial diseases are obtained by coupling CRM<sub>197</sub> to glycans from these pathogens. Wild-type DT has been described in two oligomeric forms: a monomer and a domain-swapped dimer. Their proportions depend on the chemical conditions and especially the pH, with a large kinetic barrier to interconversion. A similar situation occurs in CRM<sub>197</sub>, where the monomer is preferred for vaccine synthesis. Despite 30 years of research and the increasing application of CRM<sub>197</sub> in conjugate vaccines, until now all of its available crystal structures have been dimeric. Here, CRM<sub>197</sub> was expressed as a soluble, intracellular protein in an <i>Escherichia coli</i> strain engineered to have an oxidative cytoplasm. The purified product, called EcoCRM, remained monomeric throughout crystallization. The structure of monomeric EcoCRM is reported at 2.0 Å resolution with the domain-swapping hinge loop (residues 379–387) in an extended, exposed conformation, similar to monomeric wild-type DT. The structure enables comparisons across expression systems and across oligomeric states, with implications for monomer–dimer interconversion and for the optimization of conjugation.</p>","PeriodicalId":7029,"journal":{"name":"Acta crystallographica. Section F, Structural biology communications","volume":null,"pages":null},"PeriodicalIF":1.1000,"publicationDate":"2023-03-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"1","resultStr":"{\"title\":\"Monomeric crystal structure of the vaccine carrier protein CRM197 and implications for vaccine development\",\"authors\":\"D. Travis Gallagher, Natalia Oganesyan, Andrew Lees\",\"doi\":\"10.1107/S2053230X23002364\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p>CRM<sub>197</sub> is a genetically detoxified mutant of diphtheria toxin (DT) that is widely used as a carrier protein in conjugate vaccines. Protective immune responses to several bacterial diseases are obtained by coupling CRM<sub>197</sub> to glycans from these pathogens. Wild-type DT has been described in two oligomeric forms: a monomer and a domain-swapped dimer. Their proportions depend on the chemical conditions and especially the pH, with a large kinetic barrier to interconversion. A similar situation occurs in CRM<sub>197</sub>, where the monomer is preferred for vaccine synthesis. Despite 30 years of research and the increasing application of CRM<sub>197</sub> in conjugate vaccines, until now all of its available crystal structures have been dimeric. Here, CRM<sub>197</sub> was expressed as a soluble, intracellular protein in an <i>Escherichia coli</i> strain engineered to have an oxidative cytoplasm. The purified product, called EcoCRM, remained monomeric throughout crystallization. The structure of monomeric EcoCRM is reported at 2.0 Å resolution with the domain-swapping hinge loop (residues 379–387) in an extended, exposed conformation, similar to monomeric wild-type DT. The structure enables comparisons across expression systems and across oligomeric states, with implications for monomer–dimer interconversion and for the optimization of conjugation.</p>\",\"PeriodicalId\":7029,\"journal\":{\"name\":\"Acta crystallographica. Section F, Structural biology communications\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":1.1000,\"publicationDate\":\"2023-03-30\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"1\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Acta crystallographica. Section F, Structural biology communications\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://onlinelibrary.wiley.com/doi/10.1107/S2053230X23002364\",\"RegionNum\":4,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q4\",\"JCRName\":\"BIOCHEMICAL RESEARCH METHODS\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Acta crystallographica. Section F, Structural biology communications","FirstCategoryId":"99","ListUrlMain":"https://onlinelibrary.wiley.com/doi/10.1107/S2053230X23002364","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q4","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}
Monomeric crystal structure of the vaccine carrier protein CRM197 and implications for vaccine development
CRM197 is a genetically detoxified mutant of diphtheria toxin (DT) that is widely used as a carrier protein in conjugate vaccines. Protective immune responses to several bacterial diseases are obtained by coupling CRM197 to glycans from these pathogens. Wild-type DT has been described in two oligomeric forms: a monomer and a domain-swapped dimer. Their proportions depend on the chemical conditions and especially the pH, with a large kinetic barrier to interconversion. A similar situation occurs in CRM197, where the monomer is preferred for vaccine synthesis. Despite 30 years of research and the increasing application of CRM197 in conjugate vaccines, until now all of its available crystal structures have been dimeric. Here, CRM197 was expressed as a soluble, intracellular protein in an Escherichia coli strain engineered to have an oxidative cytoplasm. The purified product, called EcoCRM, remained monomeric throughout crystallization. The structure of monomeric EcoCRM is reported at 2.0 Å resolution with the domain-swapping hinge loop (residues 379–387) in an extended, exposed conformation, similar to monomeric wild-type DT. The structure enables comparisons across expression systems and across oligomeric states, with implications for monomer–dimer interconversion and for the optimization of conjugation.
期刊介绍:
Acta Crystallographica Section F is a rapid structural biology communications journal.
Articles on any aspect of structural biology, including structures determined using high-throughput methods or from iterative studies such as those used in the pharmaceutical industry, are welcomed by the journal.
The journal offers the option of open access, and all communications benefit from unlimited free use of colour illustrations and no page charges. Authors are encouraged to submit multimedia content for publication with their articles.
Acta Cryst. F has a dedicated online tool called publBio that is designed to make the preparation and submission of articles easier for authors.