厚壳贻贝whirlin类贝壳基质蛋白的重组表达与功能分析

Recombinant Expression and Functional Analysis of a Whirlin-like Protein from the Shell of Mytilus coruscus

贝壳历来是生物工程和材料学研究的重要对象。贝壳中的贝壳基质蛋白质在贝壳的形成与发育过程中具有重要的调控作用。Whirlin类蛋白质(Whirlin-like protein,WLP)是一种从厚壳贻贝(Mytilus coruscus)中鉴定的新型贝壳基质蛋白质。序列分析结果显示,该蛋白质含有PDZ(postsynaptic density/Discs large/Zonula occludens)结构域,而该结构域对贝壳生物矿化的影响目前尚无报道。为深入了解WLP在贝壳形成中对碳酸钙晶体的影响,在序列分析基础上,采用密码子优化结合原核重组表达,获得其重组表达产物后,开展了重组WLP对碳酸钙晶体形貌及晶型的影响研究,结晶速度抑制以及碳酸钙晶体结合分析。分析结果表明,重组WLP能诱导文石型碳酸钙晶体的形貌和方解石型碳酸钙晶体的晶型发生改变;同时重组WLP对碳酸钙晶体具有结合作用,且能抑制碳酸钙晶体的结晶速度。上述结果表明,WLP对贝壳的形成及发育具有重要影响,并可能在贝壳肌棱柱层的形成中发挥了重要作用。

As a prominent material for biomaterials and bioengineering study,the mollusk shell is composed mainly by calcium carbonate crystal and produced via the mineralization process tightly controlled by shell matrix proteins(SMPs).SMPs play an important role in the formation of the shell,as well as the regulation of calcium carbonate biomineralization and the promotion of bone growth.A novel PDZ(postsynaptic density/discs large/zonula occludens)domain-containing protein has been characterized from the myostracum layer of the shell of Mytilus coruscus previously,and was designated as whirlin-like protein(WLP).However,the roles of the PDZ domain in shell formation is unclear and no research has been reported for now.For understanding the mechanism of the PDZ domain-containing protein in biomineralization,the WLP of M.coruscus was expressed in the E.coli BL21 strain with codon optimization.The function of recombinant WLP was studied and the results showed that recombinant WLP affects the crystal morphology of aragonite calcium carbonate,as well as the polymorph of calcite calcium carbonate.Furthermore,the recombinant WLP interacts with the calcium carbonate crystal and inhibits the crystalline growth velocity of both calcite and aragonite.These results suggested that WLP may be a biomineralization-related protein in the shell of M.coruscus and play an important role in the formation of the myostracum layer.