Spinning sugars in antigen biosynthesis: characterization of the Coxiella burnetii and Streptomyces griseus TDP-sugar epimerases.
Spinning sugars in antigen biosynthesis: characterization of the Coxiella burnetii and Streptomyces griseus TDP-sugar epimerases.
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DOI：
10.1016/j.jbc.2022.101903
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发表时间：
2022-05
影响因子：
4.8
通讯作者：
Harmer, Nicholas J.
中科院分区：
文献类型：
--
作者：
Cross, Alice R.;Roy, Sumita;Vega, Mirella Vivoli;Rejzek, Martin;Nepogodiev, Sergey A.;Cliff, Matthew;Salmon, Debbie;Isupov, Michail N.;Field, Robert A.;Prior, Joann L.;Harmer, Nicholas J.
链糖和二氢羟基链糖（DHHS）分别是灰色链霉菌和伯氏柯克斯体所特有的。链糖形成抗生素链霉素的中心部分，而 DHHS 存在于人畜共患病原体伯氏梭菌的 O 抗原中。这些糖的生物合成被认为遵循与 TDP-鼠李糖相似的途径，由酶 RmlA、RmlB、RmlC 和 RmlD 催化，但确切的机制尚不清楚。链糖和 DHHS 生物合成通常需要环收缩步骤，该步骤可由 RmlC 或 RmlD 的直向同源物执行。 S. griseus 和 C. burnetii 的基因组测序已将 StrM 和 CBU1838 蛋白鉴定为这些各自物种中的 RmlC 直向同源物。在这里，我们证明这两种酶都可以执行支持 TDP-鼠李糖体内生物合成所需的 RmlC 3'',5'' 双差向异构化活性。这与在双差向异构底物上进行的环收缩步骤一致。我们进一步证明，与之前研究的直系同源物相比，3''-位置的质子交换比 5''-位置更快。我们还解析了 CBU1838 和 StrM 与 TDP 复合物的晶体结构，并表明它们形成的活性位点与之前表征的酶 RmlC、EvaD 和 ChmJ 的活性位点高度相似。这些结果支持这样的假设：链糖和DHHS是使用TDP途径生物合成的，并且RmlD旁系同源物最有可能在双差向异构化后进行环收缩。这项工作将支持阐明这些独特糖生物合成的完整途径。
The sugars streptose and dihydrohydroxystreptose (DHHS) are unique to the bacteria Streptomyces griseus and Coxiella burnetii, respectively. Streptose forms the central moiety of the antibiotic streptomycin, while DHHS is found in the O-antigen of the zoonotic pathogen C. burnetii. Biosynthesis of these sugars has been proposed to follow a similar path to that of TDP-rhamnose, catalyzed by the enzymes RmlA, RmlB, RmlC, and RmlD, but the exact mechanism is unclear. Streptose and DHHS biosynthesis unusually requires a ring contraction step that could be performed by orthologs of RmlC or RmlD. Genome sequencing of S. griseus and C. burnetii has identified StrM and CBU1838 proteins as RmlC orthologs in these respective species. Here, we demonstrate that both enzymes can perform the RmlC 3’’,5’’ double epimerization activity necessary to support TDP-rhamnose biosynthesis in vivo. This is consistent with the ring contraction step being performed on a double epimerized substrate. We further demonstrate that proton exchange is faster at the 3’’-position than the 5’’-position, in contrast to a previously studied ortholog. We additionally solved the crystal structures of CBU1838 and StrM in complex with TDP and show that they form an active site highly similar to those of the previously characterized enzymes RmlC, EvaD, and ChmJ. These results support the hypothesis that streptose and DHHS are biosynthesized using the TDP pathway and that an RmlD paralog most likely performs ring contraction following double epimerization. This work will support the elucidation of the full pathways for biosynthesis of these unique sugars.
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