Development of fine micro analytical method of drug recognition function of sugar chains

建立了一种新的毛细管电泳-质谱分析系统，用于研究糖蛋白结构对药物结合亲和力的影响。以α1-酸性糖蛋白（AGP）为模型糖蛋白，评价了AGP糖型和遗传变异体与模型基础药物的结合亲和力。(a)双触角糖链仅位于AGP分子的药物结合位点附近，对AGP与模型碱性药物（丙吡胺和维拉帕米）的对映选择性结合没有显著影响。(b)AGP有三种主要的遗传变异，A-变异，F1-变异和S-变异。A-变异体与模型药物对映体结合的对映选择性高于F1^*S变异体，而这些遗传变异体与维拉帕米对映体结合的对映选择性几乎相同。(c)采用无鞘电喷雾毛细管电泳/质谱联用技术，对胰蛋白酶AGP变异株的检测灵敏度可提高一个数量级 ...更多信息 为了研究糖蛋白的微观异质性对蛋白质功能和二级结构的影响，本文采用核磁共振、分子轨道法和毛细管电泳等方法，对钠通道蛋白和朊病毒蛋白的几个合成肽段进行了结构化学研究。合成的片段对应于负责蛋白质二级结构和/或功能的主要肽部分。发现了以下内容。(a)由于溶剂的性质和组成，模型肽片段的二级结构随环境的疏水性而变化。(b)含有疏水部分和疏水部分的聚糖产生局部疏水空间，其具有不同的微介电常数，这取决于聚糖的性质、序列和分支结构。这表明疏水局部环境的产生会引起蛋白质二级结构的改变，从而导致功能的调节和淀粉样纤维的形成。少

A novel analytical system using capillary electrophoresis (CE) and mass spectroscopy (MS) was developed to investigate the structural effect of glycoproteins upon drug binding affinity. α1-acid glycoprotein (AGP) was used as a model glycoprotein, and the binding affinities of AGP glycoforms and genetic variants to model basic drugs were evaluated.(a) The biantennary glycochains which locate only near the drug binding site on AGP molecule does not give any significant effect upon the enantioselective binding of AGP to model basic drugs (disopyramide and verapamil).(b) AGP has three major genetic variants, A-variant, F1-variant and S-variant. A-variant shows higher enantioselectivity in the binding with the enantiomers of model drugs than F1^*S variants, while these genetic variants show almost the same enantioselectivity in the binding to verapamil enantiomers.(c) CE/MS system using sheathless ESI method allows one-order higher detectability in the analyses of tryptic digests of AGP var … More iants than conventional CE method using UV detection.In order to investigate the effect of microheterogeneity of glycoproteins upon the function and the secondary structure of the proteins, the structural chemistry of several synthesized peptide fragments of Na-channel protein and prion protein were studied by using NMR, molecular orbital method and CE spectroscopy. The synthesized fragments correspond to the major peptide portion responsible for the secondary structure and/or the function of the proteins. The followings were found.(a) The secondary structure of model peptide fragments changes depending on the hydrophobicity of environment due to the nature and the component of solvents.(b) Glycans containing both hydrophobic moiety and hydrophobic moiety create local hydrophobic space with different micro-permittivity depending on nature, sequence and the branching structure of glycans. It is suggested that the creation of hydrophobic local environment would cause the change of secondary structure of protein, resulting in the regulation of function and amyloid fibril formation. Less