贻贝足的共聚焦显微拉曼光谱分析

贻贝对各种海洋环境具有广泛适应性,范围从浅海跨越到深海热液、冷泉等极端环境。贻贝主要通过其足组织分泌的蛋白形成足丝附着于岩石等固体表面,这种蛋白是一种可再生、不受水环境影响的性能良好的天然生物胶黏剂,得益于分泌蛋白的性质,足丝在水下具有黏附性强、韧性高、耐水性优良等特性,在生物材料学、医学等方面具有很好的开发潜力和应用前景,已经是国内外研究热点之一。拉曼光谱是一种非接触的、无损的可以提供分子生物化学信息的检测技术。足丝是贻贝足腺体的外在表达形式,结合扫描电镜和共聚焦显微拉曼光谱技术,从贻贝足丝的表观差异到贻贝足腺体的分泌蛋白组分和分布特征,基于深海和浅海贻贝足丝的扫描电镜表征的表观形态差异,对两种贻贝足组织分别进行共聚焦显微拉曼光谱检测,得到两种贻贝的3个腺体的拉曼光谱和腺体局部区域的2D拉曼彩色分布图,从外在表现形式足丝到内部足腺体分布,通过对比两种贻贝的3个腺体的成分以及相对分布,分析造成两种贻贝足丝差异的内在腺体分布情况,此外结合两种贻贝生存环境的差异,认为贻贝的足丝外观差异以及其内部腺体分布是贻贝应对浅海和深海冷泉完全不同理化环境的一种环境适应机制。基于实验结果得到如下结论,拉曼光谱表明两种贻贝足腺体组成:表征核心腺体的amideⅢ信号位于1 242和1 269 cm^-1位置的2个峰的峰强度相对其他两个峰(1 318和1 337 cm^-1)较高,表现为有序高级的蛋白构象,外皮和粘附盘腺体含有丰富的酪氨酸(643, 830, 850和1 615 cm^-1)和3, 4-二羟基苯丙氨酸(多巴, DOPA,785 cm^-1);浅海贻贝在1 043 cm^-1位置有高强度的胶原蛋白信号。拉曼成像呈现两种贻贝腺体分布特征:深海贻贝表现为较为集中的腺体分布,浅海贻贝腺体分布较为分散,表明贻贝为适应不同环境形成不同的腺体分布机制。由此可见,拉曼光谱可以用于研究不同环境下生存的贻贝的足腺体分布特性,并在生物样品微观分析中更多的应用。

Confocal Raman Micro-Spectroscopy Analysis of Mussel Foot

Mussels are widely distributed in various marine environments, from shallow seas to cold seeps and hydrothermal vents in the deep oceans, because of its excellent adaptabilityin different physicochemical conditions. Mussels anchor themselves on the tough substrate exposed on the seabed, such authigenic carbonate crusts or rock outcrops with adhesive protein secreted by mussel foot. This protein is a natural renewable biological adhesive available in various marine environments and mussel byssus presents such as high adhesive, high toughness and resistance to water. The adhesive protein has great potential and application prospect in biotechnology and medicine field, and it has become a research hotspot nowadays. Raman spectroscopy is a kind of non-destructive and non-invasive detection technique which can provide molecular biochemical information of organisms. In this paper, SEM and confocal Raman micro-spectroscopy are used to investigatethe apparent difference of mussel foot byssus, the secreted protein composition and distribution characteristics of mussel foot gland. Based on the apparent morphological differences of deep-sea mussels and sallow-sea mussels by SEM, two kind of mussels foot tissues are detected with confocal Raman micro-spectroscopy and acquire Raman spectra and 2 D Raman color-coded image of three glands. The components and relative distribution of three glands of two mussels are compared and byssus differences caused by the glands distribution are analyzed. Meanwhile, considering the living environments of the two kind of mussels, we think that the mussel byssus appearance difference and foot glands distribution is a kind of adaptation mechanism to the environment. The Raman spectra suggest the components of two mussels foot glands: the higher intensity at 1 242 and 1 269 cm^-1to the other two peaks(1 318 and 1 337 cm^-1) of the amide Ⅲ signals indicates that the core gland is a higher degree of conformational order protein. The proteins that comprise theplaque and cuticle enrich amino acid tyrosine(Tyr) at 643, 830, 850 and 1 615 cm^-1 and post-translationally converted to 3,4-dihydroxyphenylalanine(DOPA) at 785 cm^-1;shallow-sea mussels present high intensity collagen protein at 1 043 cm^-1. Raman imaging shows the glands distribution feature: deep-sea mussels foot glands distribution concentration and shallow-sea mussels foot glands distribution dispersion, which suggests that the glands distribution may be a kind of mechanism for the mussels to adapt different environments. The results indicate that confocal Raman micro-spectroscopy can be used in the analysis of the mussel foot glands distribution characteristics in different environments, and has great prospect to be applied in the micro-analysis of the biological samples.