García E, Arrecubieta C, Muñoz R, Mollerach M, López R
Centro de Investigaciones Biológicas (CSIC), Madrid, Spain.
Microb Drug Resist. 1997 Spring;3(1):73-88. doi: 10.1089/mdr.1997.3.73.
Type 3 pneumococci produce a capsule composed of cellobiuronic acid units connected in a beta (1-->3) linkage. Cellobiuronic acid is a disaccharide consisting of D-glucuronic acid (GlcA) beta (1-->4) linked to D-glucose (Glc). The genes implicated in the biosynthesis of the type 3 capsule have been cloned, expressed, and biochemically characterized. The three type 3-specific genes--designated as cap3ABC--are transcribed together. However, the two complete open reading frames located upstream of cap3A are not transcribed and, consequently, are not required for capsule formation. The promoter of the cap3 operon was localized by primer extension analysis. The products of cap3A, cap3B, and cap3C were biochemically characterized as a UDP-Glc dehydrogenase, the type 3 polysaccharide synthase, and a Glc-1-P uridyltransferase, respectively. The Cap3B synthase was expressed in Escherichia coli, and pneumococcal type 3 polysaccharide was synthesized in this heterologous system. When a recombinant plasmid (pLSE3B) containing cap3B was introduced by transformation into encapsulated pneumococci of types 1, 2, 5, or 8, the lincomycin-resistant transformants displayed a binary type of capsule, this is, they showed a type 3 capsule in addition to that of the recipient type. Unencapsulated (S2) laboratory strains of S. pneumoniae also synthesized a type 3 capsule when transformed with pLSE3B. On the other hand, we have cloned and sequenced seven type 1-specific genes (designated as cap1A-G), and their functions have been preliminarily assigned based on sequence similarities.
3型肺炎球菌产生一种由通过β（1→3）键连接的纤维二糖醛酸单元组成的荚膜。纤维二糖醛酸是一种由D-葡萄糖醛酸（GlcA）β（1→4）连接到D-葡萄糖（Glc）组成的二糖。与3型荚膜生物合成相关的基因已被克隆、表达并进行了生化特性分析。三个3型特异性基因——命名为cap3ABC——一起转录。然而，位于cap3A上游的两个完整开放阅读框不转录，因此，荚膜形成不需要它们。通过引物延伸分析确定了cap3操纵子的启动子。cap3A、cap3B和cap3C的产物经生化鉴定分别为UDP-葡萄糖脱氢酶、3型多糖合酶和葡萄糖-1-磷酸尿苷转移酶。Cap3B合酶在大肠杆菌中表达，并在这个异源系统中合成了肺炎球菌3型多糖。当通过转化将含有cap3B的重组质粒（pLSE3B）引入1、2、5或8型的有荚膜肺炎球菌时，耐林可霉素的转化子表现出二元型荚膜，也就是说，除了受体类型的荚膜外还显示出3型荚膜。无荚膜的（S2）肺炎链球菌实验室菌株在用pLSE3B转化时也合成了3型荚膜。另一方面，我们已经克隆并测序了七个1型特异性基因（命名为cap1A-G），并根据序列相似性初步确定了它们的功能。
