ROBUST PROTEIN FROM MUSSEL BYSSUS

贻贝粗丝状组织是一种特殊的非细胞结缔组织。它们由胶原蛋白组成，可以在几分钟内制成，在角质层的保护下可以抵抗降解，并且具有一系列令人满意的机械性能，类似于重要的生物聚合物，如肌腱胶原蛋白、弹性蛋白和拖丝。本研究的长期目标是发现足跖骨的力学性能是如何由其分子结构决定的。具体来说，提出的研究试图发现三种天然存在的融合蛋白在大丝线中的分布对这些结构在屈服后恢复初始长度和模量的能力有什么影响。这些蛋白，preCol-NG， preCol-P和preCol-D，都具有嵌段共聚物结构，由夹在两个侧结构域之间的中心胶原结构域组成，并由富含组氨酸的氨基端和羧基端覆盖。两侧结构域类似于弹性蛋白（precolp）和蜘蛛拖丝（precold）。本研究的主要目的（方法）是确定a)在纤维组装过程中前体纤维是如何定向和排列的（使用可逆双功能交联剂），b)足丝的成熟对N-和c -末端多巴残基以及前体纤维中二硫化物的形成有什么影响（苯肼衍生多巴，随后进行有限消化和MALDI TOF分析）。c)过渡金属是否通过其富含组氨酸的末端起到交联前cols的作用（通过螯合去除金属，然后进行微力学分析）；d)单个预冷分子能否在原子力显微镜下被拉伸。这些结果应该与具有广泛机械性能的脚手架生物材料的设计特别相关。

Mussel byssal threads are an extraordinary type of acellular connective tissue. Composed of collagen, they are made in minutes, resist degradation when protected by a cuticle, and have a range of desirable mechanical properties that resemble those of important biopolymers such as tendon collagen, elastin, and dragline silk. The long range objective of this research has been to discover how the mechanical properties of byssus are determined by its molecular architecture. Specifically, the proposed research attempts to discover what effect the distribution of three naturally occurring fusion proteins in byssal threads has on the ability of these structures to recover both initial length and modulus after yield.The proteins, preCol-NG, preCol-P and preCol-D, are all characterized by a block copolymer structure that consists of a central collagenous domain sandwiched between two flanking domains, and capped by histidine-rich amino- and carboxy-termini. The flanking domains resemble elastin (preCol-P) and spider dragline silk (preCol-D). The major aims (methods) of this proposal are to determine a) how preCols are oriented and aligned during fiber assembly (using reversible bifunctional crosslinkers), b) what effect maturation of byssus has on N- and C-terminal Dopa residues and on disulfide formation in preCols (phenylhydrazine derivatization for Dopa, followed by limited digestion and MALDI TOF analysis), c) whether transition metals function to cross-link preCols through their histidine-rich termini (metal removal by chelation followed by micromechanical analysis);, and d) whether individual preCol molecules can be stretched on an atomic force microscope. The outcome of these should be of particular relevance to the design of scaffolding biomaterials with a wide range of mechanical properties.