CpsE from type 2 Streptococcus pneumoniae catalyzes the reversible addition of glucose-1-phosphate to a polyprenyl phosphate acceptor, initiating type 2 capsule repeat unit formation
CpsE from type 2 Streptococcus pneumoniae catalyzes the reversible addition of glucose-1-phosphate to a polyprenyl phosphate acceptor, initiating type 2 capsule repeat unit formation
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DOI：
10.1128/jb.187.21.7425-7433.2005
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发表时间：
2005-11-01
影响因子：
3.2
通讯作者：
Yother, J
中科院分区：
文献类型：
--
作者：
Cartee, RT;Forsee, WT;Yother, J
革兰氏阳性病原体肺炎链球菌产生的 90 种荚膜类型中的大多数是通过类似于 Wzy 依赖性 O 抗原和革兰氏阴性细菌荚膜所利用的块型机制组装的。在此机制中，重复单元形成的起始（通过将糖转移至脂质受体而发生）。在肺炎链球菌中，此步骤由 CpsE（一种在大多数荚膜类型中保守的蛋白质）催化。来自肺炎链球菌 2 型菌株 D39 和含有重组 Cps2E 的大肠杆菌的膜催化将 [C-14]Glc 从 UDP-[C-14]Gic 掺入到来自两种膜的 Cps2E 依赖性糖脂产物对温和的酸水解敏感，表明 Cps2E 催化聚异戊二烯焦磷酸 Glc 的形成，向 D39 和大肠杆菌膜添加大小范围为 35 至 105 个碳的外源聚异戊二烯磷酸可刺激 Cps2E 活性。在不存在外源聚异戊二烯磷酸酯的情况下合成的糖脂产物与60碳聚异戊二烯焦磷酸Gic一起添加到含有D39膜的反应混合物中时，Cps2E活性分别被抑制40％和80％，其作为竞争性抑制剂。 UDP-Glc 也刺激 Cps2E 催化逆反应，从聚异戊二烯基焦磷酸 Gic 合成 UDP-Glc。这些数据表明，Cps2E 催化 Glc-1-P 添加到聚异戊二烯基磷酸盐受体（可能是十一异戊二烯基磷酸盐）上。
The majority of the 90 capsule types made by the gram-positive pathogen Streptococcus pneumoniae are assembled by a block-type mechanism similar to that utilized by the Wzy-dependent O antigens and capsules of gram-negative bacteria. In this mechanism, initiation of repeat unit formation (occurs by the transfer of a sugar to a lipid acceptor. In S. pneumoniae, this step is catalyzed by CpsE, a protein conserved among the majority of capsule types. Membranes from S. pneumoniae type 2 strain D39 and Escherichia coli containing recombinant Cps2E catalyzed incorporation of [C-14]Glc from UDP-[C-14]Gic into a lipid fraction in a Cps2E-dependent manner. The Cps2E-dependent glycolipid product from both membranes was sensitive to mild acid hydrolysis, suggesting that Cps2E was catalyzing the formation of a polyprenyl pyrophosphate Glc. Addition of exogenous polyprenyl phosphates ranging in size from 35 to 105 carbons to D39 and E. coli membranes stimulated Cps2E activity. The stimulation was due, in part, to utilization of the exogenous polyprenyl phosphates as an acceptor. The glycolipid product synthesized in the absence of exogenous polyprenyl phosphates comigrated with a 60-carbon polyprenyl pyrophosphate Gic. When 10 or 100 mu M UMP was added to reaction mixtures containing D39 membranes, Cps2E activity was inhibited 40% and 80%, respectively. UMP, which acted as a competitive inhibitor of UDP-Glc, also stimulated Cps2E to catalyze the reverse reaction, with synthesis of UDP-Glc from the polyprenyl pyrophosphate Gic. These data indicated that Cps2E was catalyzing the addition of Glc-1-P to a polyprenyl phosphate acceptor, likely undecaprenyl phosphate.