Biophysical and Model Studies of NiSOD

含镍超氧化物歧化酶（NiSODs）是生物体系中清除超氧化物的一种全新机制。拟议的项目使用跨学科的方法来探索在链霉菌属中发现的NiSOD的结构和功能，链霉菌属是一种负责目前医学上可用的许多抗生素的生物体。单晶X-射线衍射，X-射线吸收光谱，振动光谱和质谱将被用来检查NiSOD的结构方面。这些技术将表征蛋白质的整体结构，确定金属位点的结构和存在于各种形式的酶（包括潜在的反应中间体或抑制剂复合物）中的双氧物质，并有助于阐明四级结构在酶功能中的作用。定点突变技术和动力学测量将用于测试特定氨基酸残基在催化位点结构中的作用，并通过测量催化速率常数来监测突变的影响。动力学方法也将被用来检查四级结构的作用和一个不寻常的产品抑制，是与过氧化物在NiSOD.Broader影响：拟议的研究还提供了一个重要的培训机会，为学生在各级（本科通过博士后）在生物化学和无机化学的接口。学生将受益于与校园化学-生物学接口培训计划的联系，与美国其他机构的科学家的合作，以及与匈牙利实验室的NSF资助的交流计划。学生将接触到生物化学和无机化学的一些技术和概念。本项目由分子生物化学和无机、生物无机有机化学项目共同资助。

Nickel-containing superoxide dismutases (NiSODs) represent a completely novel mechanism for the elimination of superoxide in biological systems. The proposed project uses an interdisciplinary approach to explore the structure and function of NiSODs found in Streptomyces species, an organism that is responsible for many of the antibiotics currently available to medicine. Single crystal x-ray diffraction, x-ray absorption spectroscopy, vibrational spectroscopy and mass spectrometry will be used to examine structural aspects of NiSOD. These techniques will characterize the overall structure of the protein, determine the structure of the metal site and the dioxygen species present in various forms of the enzyme (including potential reaction intermediates or inhibitor complexes), and help to elucidate the role of quaternary structure in the function of the enzyme. Site-directed mutagenesis techniques and kinetic measurements will be used to test the role of specific amino acid residues in the structure of the catalytic site, with the effect of mutation monitored by measurement of the catalytic rate constant. Kinetic methods will also be used to examine the role of quaternary structure and an unusual product inhibition that is associated with peroxide in NiSOD.Broader Impact: The proposed research also provides a significant training opportunity for students at all levels (undergraduate through post-doctoral) at the interface of biochemistry and inorganic chemistry. Students will benefit from the association with a Chemistry-Biology Interface training program on campus, collaborations with scientists at other institutions in the U.S, and from a NSF-funded exchange program with a laboratory in Hungary. Students will be exposed to a number of techniques and concepts from both biochemistry and inorganic chemistry. Students will be drawn from a diverse pool, and effort will be made to provide opportunities for minorities.This project is funded jointly by Molecular Biochemistry and Inorganic, Bioinorganic & Organometallic Chemistry Program.