Hanada N, Kuramitsu H K
Department of Microbiology-Immunology, Northwestern University Medical-Dental Schools, Chicago, Illinois 60611.
Infect Immun. 1988 Aug;56(8):1999-2005. doi: 10.1128/iai.56.8.1999-2005.1988.
The intact gtfC gene from Streptococcus mutans GS-5 was isolated in Escherichia coli in plasmid vector pUC18. The glucosyltransferase activity expressed by the gene synthesized both low-molecular-weight water-soluble glucan and insoluble glucan in a primer-independent manner. Purification of the enzyme by procedures that minimize proteolytic digestion yielded a purified preparation with a molecular weight of 140,000. Insertional inactivation of the gtfC gene with a streptococcal erythromycin resistance gene fragment followed by transformation of strain GS-5 suggested that the gtfC gene product was required for sucrose-dependent colonization in vitro. In addition, evidence for the presence of a third gtf gene coding for soluble glucan synthesis was obtained following the construction of mutants containing deletions of both the gtfB and gtfC genes.
变形链球菌GS-5完整的gtfC基因在大肠杆菌中通过质粒载体pUC18分离得到。该基因表达的葡糖基转移酶活性以不依赖引物的方式合成了低分子量水溶性葡聚糖和不溶性葡聚糖。通过尽量减少蛋白水解消化的程序对该酶进行纯化,得到了分子量为140,000的纯化制剂。用链球菌红霉素抗性基因片段对gtfC基因进行插入失活,随后转化GS-5菌株,这表明gtfC基因产物是体外蔗糖依赖性定植所必需的。此外,在构建同时缺失gtfB和gtfC基因的突变体后,获得了存在第三个编码可溶性葡聚糖合成的gtf基因的证据。