Structure and function of bovine colostral lactoferrin

1.乳铁蛋白的相对分子质量、糖含量和二级结构经SDS聚丙烯酰胺凝胶电泳法测得乳铁蛋白的相对分子质量为81,800±2,600。碳水化合物含量中性糖为8.8%，氨基糖为2.7%。根据CD光谱计算，螺旋含量为24%。乳铁蛋白与蓝色染料牛乳铁蛋白的相互作用表明，乳铁蛋白与固定化Cibacron Blue F3GA柱有很强的相互作用。结果表明，这种相互作用是由较强的静电力引起的。用胰酶消化乳铁蛋白，得到乳铁蛋白的胰酶片段，其相对分子质量分别为52kD和27kD。通过与固定化蓝色染料的亲和层析，发现分子量较大的片段对蓝色染料和铁都有结合能力。在固定化的抗乳铁蛋白血清柱上吸附了分子量在20~40kD的片段。糖组分组成和结构的估算糖组分通过肼分解释放，然后用离子交换层析分离成中性和酸性糖组分。通过气相色谱对它们的糖组成进行了估计。固定化内切糖苷酶也被用于裂解乳铁蛋白中的碳水化合物部分。用酶凝集素测定法测定糖链的组成和结构。

1. Molecular weight, sugar content and secondary structure of lactoferrin Molecular weight of lactoferrin was estimated to be 81,800+2,600 by SDSpolyacrylamide gel electrophoresis. Carbohydrate contents were 8,8 % for neutral sugar and 2.7 % for amino sugar. Helical content was 24 % calculated from CD spectrum.2. Interacting properties of lactoferrin with blue dye Bovine lactoferrin showed strong interaction with immobilized Cibacron Blue F3GA column. It was suggested that such interaction has been due to the strong electrostatic forces.3. Tryptic fragments of lactoferrin After digestion of lactoferrin by trypsin, fragments of which molecular weight was 52 KD and 27 KD were obtained. By the affinity chromatography with immobilized blue dye, the fragment of larger molecular weight showed the binding ability both to blue dye and iron. Onto the immobilized anti-lactoferrin serum column, the fragments of which molecular weights were from 20 to 40 KD, were adsorbed.4. Estimation of composition and structure in carbohydrate moiety Carbohydrate moieties were liberated by hydrazinolysis and then were separated into neutral and acidic sugar moieties by ion exchange chromatography. They were estimated on their sugar compositions by gas chromatography. Immobilized endoblycosidases were also used to cleave the carbohydrate moieties in lactoferrin. Enzyme lectin assay was done in order to estimate the composition and the structure of the carbohydrate chains.