Stuctural studies of triple-helical proteins

描述（由申请人提供）：胶原蛋白是体内最丰富的蛋白质，其独特的三螺旋构象和重复序列模式定义为甘氨酸每三个残基。生物物理研究建议将这种模式的（Gly-X-Y）n氨基酸序列和断裂与胶原的分子特征和高阶结构联系起来，这些特征和高阶结构直接关系到胶原的功能和病理。我们对经典三螺旋结构的研究将扩展到表征在非纤维性胶原中发现的（Gly-X-Y）n模式自然断裂的后果，例如基底膜中的IV型胶原和介导真皮-表皮附着的VII型胶原。这种断裂的长度和序列对三螺旋稳定性、折叠和构象的影响将使用模型肽进行研究。为了补充肽研究，将在两个串联三螺旋模块之间引入断裂的表达细菌产物上检查柔韧性，折叠和酶敏感性。这些研究将提供有关断裂的结构后果及其生物学作用的信息。胶原蛋白分子与高阶结构的关联对其机械和生物学功能至关重要。序列之间的关系，三螺旋结合到高阶结构的过程，和最终产品的形态将被定义。研究将进一步表征观察到的胶原蛋白肽的非特异性横向组装，并在肽序列中引入静电和疏水残基，以产生更特异性的轴向相互作用和定义的高阶结构。（Gly-X-Y）n重复中的小的自然断裂似乎使侧翼三螺旋区域脱离了寄存器，它们对自结合的影响将被研究。纤原性和非纤原性胶原蛋白中的Gly错义突变可导致多种遗传性疾病。Gly错义突变的折叠、稳定性、流体动力学和构象后果将在多肽和表达的细菌结构中表征。在非纤原性胶原中，假设错义突变干扰了通过自然断裂折叠所需的再生机制。明确胶原蛋白三螺旋分子结构的基本原理及其与高阶结构的关联将进一步加深我们对正常基质结构/功能关系的理解，并促进胶原基生物材料的发展。此外，它将为确定疾病中的细胞外基质变化和开发能够抑制癌症和骨关节炎中胶原蛋白分解的药物提供基础。

DESCRIPTION (provided by applicant): Collagen is the most abundant protein in the body, defined by its unique triple-helical conformation and repeating sequence pattern with glycine as every third residue. Biophysical studies are proposed to relate the (Gly-X-Y)n amino acid sequence and breaks in this pattern with molecular features and higher order structure of collagens, which are directly related to their function and pathology. Our studies on the classic triple helix will be extended to characterization of the consequences of natural breaks in the (Gly-X-Y)n pattern found in non-fibrillar collagens, such as type IV collagen in basement membranes and type VII collagen that mediates dermal-epidermal attachment. The effects of the length and sequence of such breaks on triple-helix stability, folding and conformation will be investigated using model peptides. To complement peptide studies, flexibility, folding and enzyme susceptibility will be examined on an expressed bacterial product where a break is introduced between two tandem triple-helix modules. These studies will provide information about the structural consequences of breaks and their biological role. The association of collagen molecules to higher order structures is essential to their mechanical and biological function. The relation between sequence, the process of triple-helix association to higher order structures, and the morphology of the final product will be defined. Studies will be carried out to further characterize non- specific lateral assembly observed for collagen peptides and to introduce electrostatic and hydrophobic residues in peptide sequences to produce more specific axial interactions and defined higher order structures. Small natural breaks in the (Gly-X-Y)n repeat appear to put flanking triple-helix regions out of register, and their impact on self-association will be investigated. Gly missense mutations in fibrillar and non-fibrillar collagen lead to a variety of hereditary diseases. The folding, stability, hydrodynamic and conformational consequences of Gly missense mutations will be characterized in peptides and in an expressed bacterial construct. In non-fibrillar collagens, it is hypothesized that missense mutations interfere with the renucleation mechanism needed to fold through natural breaks. Definition of the fundamental principles of collagen triple-helix molecular structure and association into higher order structures will further our understanding of normal matrix structure/function relationships and enhance the development of collagen-based biomaterials. In addition, it will provide a basis for defining extracellular matrix alterations in disease and for developing drugs which could inhibit the breakdown of collagens in cancer and osteoarthritis.