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Biological Spectroscopy and Crystallography Using an X-ray Free Electron Laser

使用 X 射线自由电子激光进行生物光谱学和晶体学

基本信息

批准号：
9790969
负责人：
Junko Yano
金额：
$ 37.96万
依托单位：
UNIVERSITY OF CALIF-LAWRENC BERKELEY LAB
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-09-15 至 2022-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9790969
关键词：
Active Sites Biochemical Reaction Biological Biology Catalytic Domain Charge Chemical Dynamics Chemical Structure Chemistry Collaborations Communities Complex Crystallography Cytochrome P450 Data Data Analyses Data Collection Development Diffusion Electron Transport Electrons Elements Engineering Enzymes Event Evolution Feedback Goals Heme Hydrogen Bonding Hydroxyl Radical In Situ Measurement Metals Methane Methodology Methods Nature Oxidation-Reduction Oxygen Oxygenases Photons Physiologic pulse Physiological Process Property Protein Dynamics Proteins Protocols documentation Protons Radiation induced damage Reaction Resort Ribonucleotide Reductase Roentgen Rays Sampling Signal Transduction Site Source Spectrum Analysis Speed Spottings Structural Protein Structure Synchrotrons System Techniques Temperature Time Water Width X ray diffraction analysis X ray spectroscopy X-Ray Crystallography Xray Emission Spectroscopy absorption analog biological systems catalyst chemical reaction cofactor copper oxidase cryogenics design electronic structure emission spectroscopy enzyme structure experimental study flexibility geometric structure interest metalloenzyme novel novel strategies protein structure spectroscopic data tool x-ray free-electron laser

项目摘要

Project Summary/Abstract The goal of this proposal is to understand nature's well-controlled chemistry that occurs in enzymes, by following the structural dynamics of the protein and chemical dynamics of the catalyst simultaneously. It is our goal to understand the design concepts from nature with X-ray crystallography and X-ray spectroscopy techniques using X-ray Free Electron Lasers (XFELs). Although the structure of enzymes and the chemistry at the catalytic sites have been studied intensively, an understanding of the atomic-scale chemistry requires a new approach beyond the conventional steady state X-ray crystallography and X-ray spectroscopy at cryogenic temperatures. Following the dynamic changes in the geometric and electronic structure of metallo-enzymes at ambient conditions, while overcoming the severe X-ray damage to the redox active catalytic center, is key for deriving the reaction mechanism. The intense and ultra-short femtosecond (fs) X-ray pulses from X-ray free electron lasers provide an opportunity to overcome the current limitations of room temperature data collection for biological samples at synchrotron X-ray sources. The fs X-ray pulses make it possible to acquire the signal before the sample is destroyed. The objective of this proposal is to study the protein structure and dynamics of metallo-enzymes using crystallography, as well as the chemical structure and dynamics of the catalytic complexes (charge, spin, and covalency) using spectroscopy during the reaction to understand the electron-transfer processes and elucidate the mechanism. We will design and apply a full suite of time-resolved X-ray diffraction and X-ray absorption/emission spectroscopy methods, that make use of the unique properties of the XFEL beam, to follow the reaction at room temperature. This will provide an unprecedented combination of correlated data between the protein and the co-factors, all of which are necessary for a complete understanding of structure and mechanism. Spectroscopy will include both emission and absorption spectroscopy to get a complete understanding of the time-evolution of the electronic structure, while simultaneous room temperature time-resolved X-ray crystallography would provide the changes in the geometric structure of the overall protein. The systems that will be used for developing these methodologies are some of the most important metallo-enzymes in biology that use high-valent Fe and other elements for oxygen and C-H bond activation. We will focus on non-heme enzymes such as ribonucleotide reductase (Mn/Fe and Fe/Fe) and methane mono oxygenase (Fe/Fe), and heme containing Cyt P450 systems, and functional analogs of heme-copper oxidase systems engineered into simpler proteins.

项目总结/文摘