Dr. Alexey V. Cherepanov, Prof. Dr. Harald Schwalbe
{"title":"Large Temperature-Jump and Nanosecond Hyperquenching for Time-Resolved Structural Studies","authors":"Dr. Alexey V. Cherepanov, Prof. Dr. Harald Schwalbe","doi":"10.1002/cmtd.202200050","DOIUrl":null,"url":null,"abstract":"<p>The quest for atomic structures of microsecond reaction intermediates is at the frontline of modern biochemistry. Currently, there is a clear lack of experimental methods for preparing necessary time-resolved samples. Here, we report the development of a single-turnover technique for nanosecond initiation and suspension of biomolecular reactions with kinetic resolution in the microsecond time domain. Reactions can be started by large temperature-jump or direct mixing and arrested by hyperquenching in liquid cryogen at a target temperature of 77 K. Diverse morphology of nanoscale glassy bodies feature among others thin field-of-view plane sheets that can be used for structure analyses of freeze-trapped macromolecules by transmission electron cryomicroscopy. We also report the ultra-high vacuum sublimation at 77 K – a novel method for concentrating reaction intermediates for structural studies by low-temperature techniques.</p>","PeriodicalId":72562,"journal":{"name":"Chemistry methods : new approaches to solving problems in chemistry","volume":null,"pages":null},"PeriodicalIF":6.1000,"publicationDate":"2023-01-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/cmtd.202200050","citationCount":"1","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Chemistry methods : new approaches to solving problems in chemistry","FirstCategoryId":"1085","ListUrlMain":"https://onlinelibrary.wiley.com/doi/10.1002/cmtd.202200050","RegionNum":0,"RegionCategory":null,"ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"CHEMISTRY, MULTIDISCIPLINARY","Score":null,"Total":0}
引用次数: 1
Abstract
The quest for atomic structures of microsecond reaction intermediates is at the frontline of modern biochemistry. Currently, there is a clear lack of experimental methods for preparing necessary time-resolved samples. Here, we report the development of a single-turnover technique for nanosecond initiation and suspension of biomolecular reactions with kinetic resolution in the microsecond time domain. Reactions can be started by large temperature-jump or direct mixing and arrested by hyperquenching in liquid cryogen at a target temperature of 77 K. Diverse morphology of nanoscale glassy bodies feature among others thin field-of-view plane sheets that can be used for structure analyses of freeze-trapped macromolecules by transmission electron cryomicroscopy. We also report the ultra-high vacuum sublimation at 77 K – a novel method for concentrating reaction intermediates for structural studies by low-temperature techniques.