ROBUST PROTEIN SEALANT FROM MUSSEL BYSSUS

支持身体内的结构组织，包括牙齿，骨骼，皮肤， 肌腱等经常因疾病或疾病而变得脆弱或易碎 受伤。一些胶原性天然材料，如贻贝丝线， 比最健康的肌腱更坚韧、更耐用 它们的脊椎动物同行。如此出色的表现很可能是 得益于分子设计的改进。两项建议的改进措施 在骨骼中有A.骨骼胶原与类丝素的融合和 或类似Resilin的结构域，以及B.通过以下方式完全覆盖结构 一种含有多巴蛋白和邻苯二酚氧化酶的天然密封胶。在这 提案中，3个目标已经被实验性地勾勒出来：1)丝绸-和 肌间胶原蛋白的Resilin样结构域将被表征 生物化学和标准重组DNA方法学；2)密封剂 将对邻苯二酚氧化酶进行测序，并使用免疫金进行原位定位 标记，并利用模拟自然的多肽进行动力学研究 底物；3)萌发前体物之间的分子相互作用 BYSAL线程将使用双功能可逆交叉- 接头结合酸尿素聚丙烯酰胺凝胶电泳法， 电印迹聚偏二氟乙烯膜和基质 印迹辅助激光解吸电离质谱仪 蛋白质。

Supporting structural tissues in the body including tooth, bone, skin, tendon, etc. often become weak or brittle as a result of disease or injury. Some collagenous natural materials like mussel byssal threads are significantly tougher and more durable than the healthiest tendons of their vertebrate counterparts. Such superior performance is likely to be endowed by improvements in molecular design. Two suggested improvements in byssus are A. the amalgamation of byssal collagen with fibroin-like and or resilin like domains, and B. the complete coating of the structure by a natural sealant containing a Dopa-protein and catechol oxidase. In this proposal, 3 goals have been experimentally delineated: 1) The silk- and resilin-like domains of byssal collagens will be characterized biochemically and by standard recombinant DNA methodology; 2) the sealant catechol oxidase will be sequenced, localized in situ using immunogold labelling, and studied kinetically using peptides modelled after natural substrates; 3) Molecular interactions among the precursors in nascent byssal threads will be examined using a bifunctional reversible cross- linker in combination with acid-urea polyacrylamide gel electrophoresis, electroblotting to polyvinylidene difluoride membranes, and matrix assisted laser desorption ionization mass spectrometry of blotted proteins.