New X-ray Diffractometer and Detector for Purdue Macromolecular Crystallography

摘要 通过X射线结晶学在原子分辨率下确定大分子结构可以说是 上个世纪生命科学中最伟大的技术成就。通过让分子发光 这项技术极大地加深了我们对健康和疾病的理解 加快开发更具选择性、更有效、更安全的药物。蛋白质的故事 普渡大学的结晶学始于1964年，当时聘请了该领域的先驱迈克尔·罗斯曼。一个 正式的生物物理学培训计划于1989年启动，并获得了NRSA机构博士前培训补助金， 与此同时，马基基金会获得了数百万美元的赠款，该基金为招聘提供了资金 现有X射线设施中现有的额外教员和X射线发生器。装备精良的设施 和致力于推进分子生物物理学和初级科学家培训的专家教员 产生了许多成功的调查人员，包括吴浩(哈佛)，梁彤(哥伦比亚)，马文·哈克特 (德克萨斯大学奥斯汀分校)和杰克·约翰逊(斯克里普斯大学)。约翰·特斯默，这项提议的PI，同样也是普渡大学的实习生。 普渡也是该领域创新的重要枢纽。例如，罗斯曼博士和罗斯曼博士。 珍妮特·史密斯分别是分子置换和MAD相变技术的关键贡献者， 这是目前世界上最常用的两种定相技术。普渡结构的新纪元 生物学已经开始了。2009年，该大学开设了霍克迈尔结构生物学大厅，这是一个耗资3000万美元的设施 以及大分子晶体学核心的新家。普渡重申其对结构性改革的承诺 生物通过2019年购买RockImager Duo水晶酒店和成像系统来更新核心， 和一台牛津冷冻系统800系列液氮冷却器。重新启动我们的T32分子生物物理培训 该计划(2019-24年资助)现在每年支持7名实习生(Tesmer PI)。这项建议的具体目的是 通过更换我们老化的~30年的、无法再维修的X射线发生器来满足关键需求 一种与同步加速器耦合的现代、可靠且经济实惠的MicroMax-007高频旋转阳极发电机 高质量，Dectris Eiger2混合光子计数探测器。发电机及其先进的光学系统将提供一种 晶体上的90微米X射线光束非常明亮，可优化小样品的信噪比。大型的155x163 毫米探测器将是微弱反射的理想选择，具有小的75微米像素，可以分辨近距离的反射。A 4- 圆形卡帕测角仪将增加数据收集的灵活性。新的设置将使X射线通量提高100倍， 探测器的灵敏度大约是成像板的10倍，从而提高了我们的整体 性能提高了大约三个数量级，并能够从 即使是最具挑战性的水晶。新的格局将极大地造福于科学和教育 至少20名教员的使命，不仅为他们的目标提供高分辨率的生物学见解，而且还 加快开发针对癌症和神经疾病的新疗法。

Summary Macromolecular structure determination at atomic resolution via X-ray crystallography is arguably one of the greatest technical achievements in the life sciences over the last century. By shedding light on molecular mechanisms essential for life, the technique has dramatically deepened our understanding of health and disease and accelerated the development of drugs that are more selective, potent, and safe. The story of protein crystallography at Purdue began in 1964 with the recruitment of Michael Rossmann, a pioneer in the field. A formal biophysics training program was instituted in 1989 with an NRSA Institutional Predoctoral Training Grant, coinciding with the award of a multi-million dollar Markey Foundation grant, which provided funds for hiring additional faculty and the X-ray generators currently in the existing X-ray facility. Armed with excellent facilities and expert faculty dedicated to advancing molecular biophysics and the training of junior scientists, Purdue produced many successful investigators including Hao Wu (Harvard), Liang Tong (Columbia), Marvin Hackert (UT Austin), and Jack Johnson (Scripps). John Tesmer, the PI of this proposal, is likewise a Purdue trainee. Purdue has also served as an important hub for innovation in the field. For example, Dr. Rossmann and Dr. Janet Smith were key contributors to the techniques of molecular replacement and MAD phasing, respectively, which are now the two most commonly used phasing techniques world-wide. A new era in Purdue structural biology has begun. In 2009, the university opened Hockmeyer Hall of Structural Biology, a 30 million dollar facility and the new home of the Macromolecular Crystallography core. Purdue reaffirmed its commitment to structural biology by updating the core with the 2019 purchase of a RockImager Duo crystal hotel and imaging system, and an Oxford Cryosystems 800 Series liquid nitrogen cooler. A reboot of our T32 Molecular Biophysics Training Program (funded 2019-24) now supports 7 trainees per year (Tesmer PI). The specific aim of this proposal is to fulfill a critical need by replacing our aging ~30 year old X-ray generators, which can no longer be repaired, with a modern, reliable, and cost effective MicroMax-007 HF rotating anode generator coupled to a synchrotron quality, Dectris Eiger2 hybrid photon counting detector. The generator and its advanced optics will provide a brilliant, small 90 µm X-ray beam at the crystal, optimizing signal-to-noise for small samples. The large 155x163 mm detector will be ideal for weak reflections, with small 75 µm pixels to resolve closely spaced reflections. A 4- circle kappa goniometer will add flexibility in data collection. The new setup will improve X-ray flux >100-fold, and the detector sensitivity is about 10 times greater than imaging plates, thereby improving our overall performance by about three orders of magnitude and enabling the collection of superior diffraction data from even the most challenging crystals. The new configuration will greatly benefit the scientific and educational mission of at least 20 faculty by not only providing high resolution biological insights into their targets, but also accelerating the development of novel therapeutics targeting cancer and neurological disease.