{"title":"膜内GlpG蛋白水解的时间分辨结构快照","authors":"Sangwoo Cho, R. Baker, Ming Ji, S. Urban","doi":"10.2210/pdb6pj8/pdb","DOIUrl":null,"url":null,"abstract":"Protein cleavage inside the cell membrane triggers various pathophysiological signaling pathways, but the mechanism of catalysis is poorly understood. We solved ten structures of the Escherichia coli rhomboid protease in a bicelle membrane undergoing time-resolved steps that encompass the entire proteolytic reaction on a transmembrane substrate and an aldehyde inhibitor. Extensive gate opening accompanied substrate, but not inhibitor, binding, revealing that substrates and inhibitors take different paths to the active site. Catalysis unexpectedly commenced with, and was guided through subsequent catalytic steps by, motions of an extracellular loop, with local contributions from active site residues. We even captured the elusive tetrahedral intermediate that is uncleaved but covalently attached to the catalytic serine, about which the substrate was forced to bend dramatically. This unexpectedly stable intermediate indicates rhomboid catalysis uses an unprecedented reaction coordinate that may involve mechanically stressing the peptide bond, and could be selectively targeted by inhibitors. Time-resolved crystallography captures Escherichia coli rhomboid protease GlpG in different steps of catalysis, revealing how substrate reaches the active site and reaction intermediates.","PeriodicalId":18836,"journal":{"name":"Nature Structural &Molecular Biology","volume":null,"pages":null},"PeriodicalIF":16.8000,"publicationDate":"2019-09-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"3","resultStr":"{\"title\":\"Time-resolved structural snapshot of proteolysis by GlpG inside the membrane\",\"authors\":\"Sangwoo Cho, R. Baker, Ming Ji, S. Urban\",\"doi\":\"10.2210/pdb6pj8/pdb\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"Protein cleavage inside the cell membrane triggers various pathophysiological signaling pathways, but the mechanism of catalysis is poorly understood. We solved ten structures of the Escherichia coli rhomboid protease in a bicelle membrane undergoing time-resolved steps that encompass the entire proteolytic reaction on a transmembrane substrate and an aldehyde inhibitor. Extensive gate opening accompanied substrate, but not inhibitor, binding, revealing that substrates and inhibitors take different paths to the active site. Catalysis unexpectedly commenced with, and was guided through subsequent catalytic steps by, motions of an extracellular loop, with local contributions from active site residues. We even captured the elusive tetrahedral intermediate that is uncleaved but covalently attached to the catalytic serine, about which the substrate was forced to bend dramatically. This unexpectedly stable intermediate indicates rhomboid catalysis uses an unprecedented reaction coordinate that may involve mechanically stressing the peptide bond, and could be selectively targeted by inhibitors. Time-resolved crystallography captures Escherichia coli rhomboid protease GlpG in different steps of catalysis, revealing how substrate reaches the active site and reaction intermediates.\",\"PeriodicalId\":18836,\"journal\":{\"name\":\"Nature Structural &Molecular Biology\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":16.8000,\"publicationDate\":\"2019-09-30\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"3\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Nature Structural &Molecular Biology\",\"FirstCategoryId\":\"1085\",\"ListUrlMain\":\"https://doi.org/10.2210/pdb6pj8/pdb\",\"RegionNum\":1,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"\",\"JCRName\":\"\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Nature Structural &Molecular Biology","FirstCategoryId":"1085","ListUrlMain":"https://doi.org/10.2210/pdb6pj8/pdb","RegionNum":1,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"","JCRName":"","Score":null,"Total":0}
Time-resolved structural snapshot of proteolysis by GlpG inside the membrane
Protein cleavage inside the cell membrane triggers various pathophysiological signaling pathways, but the mechanism of catalysis is poorly understood. We solved ten structures of the Escherichia coli rhomboid protease in a bicelle membrane undergoing time-resolved steps that encompass the entire proteolytic reaction on a transmembrane substrate and an aldehyde inhibitor. Extensive gate opening accompanied substrate, but not inhibitor, binding, revealing that substrates and inhibitors take different paths to the active site. Catalysis unexpectedly commenced with, and was guided through subsequent catalytic steps by, motions of an extracellular loop, with local contributions from active site residues. We even captured the elusive tetrahedral intermediate that is uncleaved but covalently attached to the catalytic serine, about which the substrate was forced to bend dramatically. This unexpectedly stable intermediate indicates rhomboid catalysis uses an unprecedented reaction coordinate that may involve mechanically stressing the peptide bond, and could be selectively targeted by inhibitors. Time-resolved crystallography captures Escherichia coli rhomboid protease GlpG in different steps of catalysis, revealing how substrate reaches the active site and reaction intermediates.
期刊介绍:
Nature Structural & Molecular Biology is a monthly journal that focuses on the functional and mechanistic understanding of how molecular components in a biological process work together. It serves as an integrated forum for structural and molecular studies. The journal places a strong emphasis on the functional and mechanistic understanding of how molecular components in a biological process work together. Some specific areas of interest include the structure and function of proteins, nucleic acids, and other macromolecules, DNA replication, repair and recombination, transcription, regulation of transcription and translation, protein folding, processing and degradation, signal transduction, and intracellular signaling.