Development of immunological method for prevention of dental caries using genetic engineering

利用基因工程开发预防龋齿的免疫学方法

• 批准号: 04557095
• 负责人: KOGA Toshihiko
• 金额: $ 6.72万
• 依托单位: KYUSHU UNIVERSITY (1993-1994)国立予防衛生研究所 (1992)
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Developmental Scientific Research (B)
• 财政年份: 1992
• 资助国家: 日本
• 起止时间: 1992 至 1994
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-04557095/
• 关键词: Dental caries; Streptococcus mutans; Adherence factor; Protein antigen; Glucosyltransferase; Fusion protein; スクロース; 付着; 免疫; 遺伝子

The first step for induction of dental caries by Streptococcus mutans is the adherence of the organism to tooth surfaces. Surface protein antigen (PAc) is considered to participatein the sucrose-independent attachment of the cell to acquired pellicles on tooth surfaces. On the other hand, glucosyltransferase (GTF) is important in the sucrose-dependent adherence of S.mutans.Our epitope mapping using many decapeptides showed that antigenic epitopes were located in the N-terminal A-repeat and middle region of PAc.Moreover, the A-repeat bound to salivary components. We have, therefore, constructed a fusion protein of the fragment corresponding to the A-repeat of PAc and the fragment corresponding to the sucrose-binding site of GTF-I,using an expression plasmid vector pTrc99A.Escherichia coli XL1-blue containing the chimeric plasmid pFPG22 expressed a fusion protein of PAc and GTF-I that reacted with both rabbit anti-PAc serum and antiGTF-I serum. Subcutaneous immunization with this fusion protein induced high titers of antibodies to both adherence factors, and these antibodies inhibited GTF-I, suggesting that the fusion protein may be useful for immunological prevention of dental caries.

变形链球菌诱导龋齿的第一步是生物体粘附到牙齿表面。表面蛋白抗原（PAc）被认为参与了细胞在牙齿表面获得性膜上的非蔗糖依赖性附着。另一方面，葡糖基转移酶（glucosyltransferase，GTF）在变形链球菌的蔗糖依赖性粘附中起重要作用，我们利用多种十肽进行抗原表位定位，发现抗原表位位于PAc的N端A重复序列和中间区域，并且A重复序列与唾液组分结合。因此，我们使用表达质粒载体pTrc 99 A构建了对应于PAc的A重复序列的片段和对应于GTF-1的蔗糖结合位点的片段的融合蛋白。含有嵌合质粒pFPG 22的大肠杆菌XL 1-blue表达了PAc和GTF-1的融合蛋白，其与兔抗PAc血清和抗GTF-1血清均反应。用该融合蛋白皮下免疫诱导高滴度的抗体的粘附因子，这些抗体抑制GTF-I，这表明融合蛋白可能是有用的免疫预防龋齿。

项目成果

K. Matsushita 他4名: "Identification of the antigenic epitopes in a surface protein antigen of Streptococcus mutans in humans" Infection and Immunity. 62. 4034-4042 (1994)

K. Matsushita 和其他 4 人：“人类变形链球菌表面蛋白抗原中抗原表位的鉴定”《感染与免疫》62. 4034-4042 (1994)。

N. Okahashi 他1名: "Pulse-field gel electrophoresis: large scale restriction maps" Methods in Gene Technology. 2. 207-226 (1994)

N. Okahashi 等人：“脉冲场凝胶电泳：大规模限制性图谱”基因技术方法 2. 207-226 (1994)。

N.Okahashi et al.: "Pulse-field gel electrophoresis : large scale restriction maps" Methods in Gene Technology. vol.2. 207-226 (1994)

N.Okahashi 等人：“脉冲场凝胶电泳：大规模限制性图谱”基因技术方法。

G.Hajishengallis: "Affinity and specificity of the interactions between Streptococcus mutans antigen I/II and salivary components" Journal of Dental Research. 73. 1493-1502 (1994)

G.Hajishengallis：“变形链球菌抗原 I/II 与唾液成分之间相互作用的亲和力和特异性”《牙科研究杂志》。

N.Okahashi: "Identification of antigenic epitopes in an alanine-rich repeating region of a surface antigen of Streptococcus mutans." Infection and Immunity. 61. 1301-1306 (1993)

N.Okahashi：“变形链球菌表面抗原富含丙氨酸的重复区域中抗原表位的鉴定。”