STRUCTURAL AND DYNAMIC STUDY OF MODEL HEME COMPLEXES

实验方法，和应用，NMR光谱的O2结合肌红蛋白和血红蛋白的特别顺磁性衍生物将进行探索和扩展，以阐明其连接和autoxidizability的电子和分子结构控制机制。 最终目的是了解氧结合的分子机制，以帮助设计基因工程血红蛋白作为血液替代品，并了解自然突变血红蛋白的病理性质。该计划将继续开发有效的方法，用于确定性分配和血红素腔的分子结构测定，并将强调开发强大的基础，用于解释每个球蛋白氧化/自旋状态在功能相关的结构决定因素方面的超精细位移。后一个目标将依赖于对一系列不同的、结构特征化的参考球蛋白、这些参考球蛋白的一系列点突变体的详细研究，这些点突变体被设计为扰乱活性位点的所选结构/动力学性质，以及参考和点突变的球蛋白，其用经修饰的血红素重建，所述经修饰的血红素被设计为将乙烯基的菱形影响与球蛋白腔的特征分离。超精细的位移在氰珠蛋白将被解释在轴向他的取向，His-Fe键强度以及远端残基氢键，和空间相互作用，结合配体。球蛋白诱导的超精细位移和轴向His方向的变化之间的相关性将被建立，探针定量部分水结扎完善，和热力学和动力学的特点，这样的水结扎。确定脱氧球蛋白中主磁轴的不寻常取向的远端相互作用的性质，以及血红素接触移位模式与His取向和非结扎远端水的存在之间的相关性将被建立。所开发的方法将用于解决摇蚊Hb中远端His的溶液与晶体取向的差异，结构表征在热变性中的平衡中间体，表征序列比对，折叠拓扑结构，并确定一系列“小型”或“紧凑”的远端残基。（109-118个残基）球蛋白，其特征是极端亲O2，有时极端抗自氧化的吸虫球蛋白中的远端氢键网络，并表征了远端相互作用和近端His-Fe键应变的变化，其伴随着连接的HbA中的变构转变。

The experimental approaches to, and application of, NMR spectroscopy of particularly paramagnetic derivatives of O2 binding myoglobin and hemoglobin will be explored and expanded to elucidate electronic and molecular structural control mechanisms of their ligation and autoxidizability. The ultimate aim is to understand the molecular mechanism of oxygen binding so as to aid in the design of genetically engineered hemoglobin as a blood substitute and to understand the pathological properties of natural mutant hemoglobins. The program continues to develop effective methods for definitive assignments and molecular structural determination of the heme cavity, and will emphasize the development of robust bases for interpreting hyperfine shifts for each of the globin oxidation/spin states in terms of functionally relevant structural determinants. This latter goal will rely on detailed studies on a series of diverse, structurally characterized reference globins, a series of point mutants of these reference globin designed to perturb selected structurally/dynamic properties of the active site, as well as both reference and point mutated globins reconstituted with modified hemes designed to separate the rhombic influences of the vinyls from that characteristic of the globin cavity. The hyperfine shifts in cyanomet globins will be interpreted in terms of the axial His orientation, the His-Fe bond strength as well as distal residue hydrogen bonding to, and steric interaction with, the bound ligand. The correlation between globin induced changes in hyperfine shift and axial His orientation in metglobins will be established, probes to quantitate partial water ligation perfected, and the thermodynamics and dynamics of such water ligation characterized. The nature of the distal interactions that determine the unusual orientation of the major magnetic axis in deoxy globins, as well as the correlation between heme contact shift pattern and both His orientation and the presence of non-ligated distal water will be established. The developed approaches will be used to resolve differences in the solution versus crystal orientation of the distal His in Chironomus Hb, structurally characterize equilibrium intermediates in the thermal denaturation of Aplysia Mb, characterize the sequence alignment, folding topology and identify distal residues in a series of "mini" or "compact" (109-118 residue) globins, characterize the distal H- bonding network in the extremely O2-avid, and on occasion, extremely autoxidative-resistant, trematode globins, and characterize the changes in distal interactions and proximal His-Fe bond strain that accompanies the allosteric transition in ligated HbA.