Jespersgaard C, Hajishengallis G, Greenway T E, Smith D J, Russell M W, Michalek S M
Departments of Microbiology, University of Alabama at Birmingham, Birmingham, Alabama 35294, USA.
Infect Immun. 1999 Feb;67(2):810-6. doi: 10.1128/IAI.67.2.810-816.1999.
Glucosyltransferase (GTF) enzymes of mutans streptococci are considered virulence factors due to their ability to synthesize adhesive glucans, which facilitate cell-to-cell adherence and accumulation. In this study we report the cloning, expression, and characterization of the catalytic (CAT) and glucan-binding (GLU) domains of S. mutans GTF-I encoded by gtfB. The CAT and GLU polypeptides represent amino acid residues 253 to 628 and 1183 to 1473, respectively, of S. mutans GTF-I. Antibodies to recombinant CAT and GLU were generated in rabbits and purified by affinity chromatography. Purified anti-CAT antibodies significantly inhibited water-insoluble glucan synthesis by S. mutans and S. sobrinus GTFs (P < 0.0001 and P < 0.05, respectively). The purified anti-GLU antibodies significantly inhibited both water-insoluble and water-soluble glucan synthesis by S. mutans GTFs (P < 0.0001 and P < 0.05, respectively). These results demonstrate that anti-CAT and anti-GLU antibodies are capable of inhibiting a variety of GTF activities. Since antibodies to S. mutans in saliva are implicated in protection against disease, we next assessed the ability of CAT and GLU polypeptides to induce mucosal antibody responses in mice. Intranasal (i.n.) immunization of mice with CAT showed significantly (P < 0.005) elevated levels of specific immunoglobulin G (IgG) antibody activity in serum and specific IgA antibody activity in serum, saliva, vaginal washes, and fecal samples. GLU immunized animals showed significantly (P < 0.005) elevated levels of specific IgA antibody activity in serum and vaginal secretions. Taken together, these results demonstrate that the recombinant CAT and GLU polypeptides are effective in inducing both mucosal and systemic immune responses. The ability of these polypeptides to induce a mucosal IgA immune response in mice after i.n. immunization supports their use as subunit vaccine candidates in the development of an anticaries vaccine.
变形链球菌的葡糖基转移酶（GTF）因其能够合成粘性葡聚糖而被视为毒力因子，这些葡聚糖有助于细胞间的粘附和聚集。在本研究中，我们报道了由gtfB编码的变形链球菌GTF-I的催化（CAT）结构域和葡聚糖结合（GLU）结构域的克隆、表达及特性分析。CAT和GLU多肽分别代表变形链球菌GTF-I的第253至628位和第1183至1473位氨基酸残基。用重组CAT和GLU免疫兔子并通过亲和层析纯化抗体。纯化的抗CAT抗体显著抑制变形链球菌和远缘链球菌GTF的水不溶性葡聚糖合成（分别为P < 0.0001和P < 0.05）。纯化的抗GLU抗体显著抑制变形链球菌GTF的水不溶性和水溶性葡聚糖合成（分别为P < 0.0001和P < 0.05）。这些结果表明抗CAT和抗GLU抗体能够抑制多种GTF活性。由于唾液中抗变形链球菌的抗体与疾病预防有关，我们接下来评估了CAT和GLU多肽在小鼠中诱导粘膜抗体反应的能力。用CAT对小鼠进行鼻内（i.n.）免疫后，血清中特异性免疫球蛋白G（IgG）抗体活性以及血清、唾液、阴道冲洗液和粪便样本中特异性IgA抗体活性显著升高（P < 0.005）。用GLU免疫的动物血清和阴道分泌物中特异性IgA抗体活性显著升高（P < 0.005）。综上所述，这些结果表明重组CAT和GLU多肽在诱导粘膜和全身免疫反应方面是有效的。这些多肽在鼻内免疫后能够在小鼠中诱导粘膜IgA免疫反应，这支持了它们作为亚单位疫苗候选物用于开发抗龋疫苗。
