Ultraviolet (UV) light microscope for protein and nucleic acid crystal detection

用于蛋白质和核酸晶体检测的紫外 (UV) 光学显微镜

• 批准号: RTI-2017-00566
• 负责人: Mark, Brian
• 金额: $ 8.06万
• 依托单位: University of Manitoba
• 依托单位国家: 加拿大
• 项目类别: Research Tools and Instruments
• 财政年份: 2016
• 资助国家: 加拿大
• 起止时间: 2016-01-01 至 2017-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=616712
• 关键词: Ultraviolet; UV; light; microscope; protein

Macromolecular X-ray crystallography provides unparalleled insight into the molecular basis of life, since it is a method that can be used to visualize the three-dimensional structure of biomolecules such as proteins and nucleic acids at or near atomic resolution. The University of Manitoba has developed a strong presence in the field over the past 10 years, which includes the research-intensive laboratories of Drs. Mark, Stetefeld and McKenna. Together, our laboratories are staffed by 29 HQP and trainees who investigate topics ranging from the structural biology of host-pathogen interactions and antibiotic resistance to matrix biology and chlorophyll biosynthesis. Key to the success of any biological X-ray crystallography research program is the ability to identify crystals of target proteins, nucleic acids, or larger quaternary complexes of these molecules in crystallization experiments. Hundreds of chemical conditions must typically be screened using a stereo microscope to identify the few conditions in which a target macromolecule or quaternary complex will crystalize. Crystals that do grow are often very small (microcrystals) and have low contrast; thus, they are easily missed when using conventional brightfield illumination techniques. Illumination with ultraviolet (UV) light, however, readily overcomes this problem since it markedly enhances the presence of protein and/or nucleic acid crystals by enabling direct visualization of the intrinsic tryptophan fluorescence and UV absorption of protein and nucleic acid crystals, respectively. This is especially useful when crystals are growing within dense precipitates that often form in crystallization experiments. Moreover, UV light microscopy can be used to distinguish crystals of protein and/or nucleic acid from those of salt, the latter of which often arise in crystallization experiments and can be highly misleading. Given the remarkable advantages of UV light microscopy, it has become a vital part of modern biological X-ray crystallography research. Unfortunately, there is no UV light microscope in the province of Manitoba that is designed to detect protein or nucleic acid crystals during crystallization experiments. This puts Manitoba structural biologists and their HQP trainees at a distinct disadvantage by restricting their ability to advance structural studies of macromolecules that yield microcrystals which are very difficult, if not impossible, to detect by conventional light microscopy. To overcome this significant limitation, we request funding for a reasonably priced UV-light microscope that will fundamentally improve the success of our crystal screening campaigns and quality of our HQP training. The microscope would integrate into our X-ray crystallography facility at the University of Manitoba and fill a crucial technology gap that is needed to sustain our research and HQP training capabilities.

大分子x射线晶体学提供了对生命分子基础的无与伦比的洞察，因为它是一种可以用于在或接近原子分辨率下可视化生物分子（如蛋白质和核酸）的三维结构的方法。在过去的10年里，马尼托巴大学在该领域发展了强大的存在，其中包括博士的研究密集型实验室。马克，斯特菲尔德和麦肯纳。我们的实验室共有29名HQP和实习生，他们研究的主题从宿主-病原体相互作用的结构生物学和抗生素耐药性到基质生物学和叶绿素生物合成。