Purification, characterization and structure analyzes of calcium binding protein of oral steptococci.

口腔链球菌钙结合蛋白的纯化、表征和结构分析。

• 批准号: 08672380
• 负责人: YAMAGUCHI Taihei
• 金额: $ 1.41万
• 依托单位: KAGOSHIMA UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08672380/
• 关键词: oral Streptococcus milleri; fimbriae; calcium dependence; saliva aggregation; calcium binding protein; saliva agglutinin; adhernce activity; 抗原物質; 局所免疫

H.8.A part of Streptococcus intermedius was recognized by human salivary agglutinin in the presence of calcium ion. In this study, a calcium binding protein has been purified from the bacterial sonicated extract of a isolate of, 1208-1, by Phenyl Sepharose HP hydrophobic column chromatography and Mono Q anion-exchange chromatography, followed by the second Mono Q column and Superose 12 gel filtration in the presence of biionic detergent, CHAPS.The purified material has a Mr of 51 kDa as homogeneic monomer meterial and approximately 200 kDa as tetramer by SDS-PAGE and by gel filtration. The isoelectric point was pH 4.7. The ^<45>Ca autoradiograph showed that only the monomer protein bound to Ca^<2+>.H.9.The saliva aggregation of 1208-1 cells was depended on amount of saliva and on calcium ion over pH 5.5. The saliva agglutinin was purified by Sephacrul S400HR gel filtration. SDS-PAGE indicated that the agglutinins were two high molecular weight proteins (-100kDa). Both saliva filtrate a … More nd the purified agglutinin boiled for 5 min showed no aggregation activity. Adsorption analyzes showed that the present agglutinins were related with aggregation of a S.intermedius K1K isolate and a Streptococcus mutans MT8148 strain also, but another mechanisim related with saliva aggregation of a S.intermedius K16-1K isolate. Adherence assay showed that the agglutinin on the surface of actinomyces cells and hydroxyapatite beads accounted for binding of streptococcus cell and that agglutinin effectively inhibited the adhernce to apatite beads coated with native saliva but not to actinomyces cells. Saliva filtrates from 10 donors showed different aggregating activity. Adherence activites of streptococcal cells to actinomyces cells and apatite beads coated with each native saliva were not related with aggregation activity.Each saliva filtrates as effector inhibited the adherence to coated apatite deads in proportion to aggregation titer. SDS-PAGE demonstrated that the depth of the agglutinin bands related with aggregative titer. Less

H.8.在钙离子存在下，人唾液凝集素可识别部分中间链球菌。本研究通过Phenyl Sepharose HP疏水柱层析和Mono Q阴离子交换层析，然后在双离子去污剂存在下通过第二次Mono Q柱和Superose 12凝胶过滤，从1208-1分离物的细菌超声提取物中纯化了钙结合蛋白，通过SDS-PAGE和凝胶过滤，纯化的材料具有作为凝胶单体材料的51 kDa的Mr和作为四聚体的约200 kDa的Mr。等电点为pH 4.7。1208 <45>-1细胞的唾液聚集依赖于唾液量和pH 5.5以上的钙离子。用Sephacrul S400 HR凝胶过滤法纯化唾液凝集素。SDS-PAGE分析表明，凝集素为两种高分子量蛋白质（~ 100 kDa）。唾液滤液和 ...更多信息 纯化的凝集素煮沸5 min后无凝集活性。吸附分析表明，该凝集素与中间型链球菌K1 K和变形链球菌MT 8148菌株的聚集有关，而与中间型链球菌K16- 1 K菌株的唾液聚集有关。粘附实验表明放线菌细胞与羟基磷灰石微球表面的凝集素是放线菌细胞与羟基磷灰石微球结合的主要原因，凝集素能有效抑制放线菌细胞与天然唾液包被的羟基磷灰石微球的粘附，但对放线菌细胞的粘附无明显影响。10个供者的唾液凝集活性不同。链球菌细胞对放线菌细胞和涂有天然唾液的磷灰石珠的粘附活性与聚集活性无关，唾液作为效应物抑制涂有天然唾液的磷灰石珠的粘附与聚集滴度成正比。SDS-PAGE显示凝集素条带的深浅与凝集效价有关。少

项目成果

山口 泰平・井上 昌一: "g血清型ミレリ連鎖球菌に対する唾液凝集因子の精製と性状" 日本細菌学雑誌. 49(1). 96-96 (1994)

Taihei Yamaguchi 和 Shoichi Inoue：“针对 G 血清型链球菌的唾液凝集因子的纯化和特性”日本细菌学杂志 49(1) (1994)。