Rodas Paula I, Álamos-Musre A Said, Álvarez Francisca P, Escobar Alejandro, Tapia Cecilia V, Osorio Eduardo, Otero Carolina, Calderón Iván L, Fuentes Juan A, Gil Fernando, Paredes-Sabja Daniel, Christodoulides Myron
Center for Integrative Medicine and Innovative Sciences, Facultad de Medicina, Universidad Andres Bello, Santiago, Chile
Center for Integrative Medicine and Innovative Sciences, Facultad de Medicina, Universidad Andres Bello, Santiago, Chile.
FEMS Microbiol Lett. 2016 Sep;363(17). doi: 10.1093/femsle/fnw181. Epub 2016 Jul 26.
The ADP-ribosylating enzymes are encoded in many pathogenic bacteria in order to affect essential functions of the host. In this study, we show that Neisseria gonorrhoeae possess a locus that corresponds to the ADP-ribosyltransferase NarE, a previously characterized enzyme in N. meningitidis The 291 bp coding sequence of gonococcal narE shares 100% identity with part of the coding sequence of the meningococcal narE gene due to a frameshift previously described, thus leading to a 49-amino-acid deletion at the N-terminus of gonococcal NarE protein. However, we found a promoter region and a GTG start codon, which allowed expression of the protein as demonstrated by RT-PCR and western blot analyses. Using a gonococcal NarE-6xHis fusion protein, we demonstrated that the gonococcal enzyme underwent auto-ADP-ribosylation but to a lower extent than meningococcal NarE. We also observed that gonoccocal NarE exhibited ADP-ribosyltransferase activity using agmatine and cell-free host proteins as ADP-ribose acceptors, but its activity was inhibited by human β-defensins. Taken together, our results showed that NarE of Neisseria gonorrhoeae is a functional enzyme that possesses key features of bacterial ADP-ribosylating enzymes.
许多致病细菌中都编码有ADP核糖基化酶,以影响宿主的基本功能。在本研究中,我们发现淋病奈瑟菌拥有一个与ADP核糖基转移酶NarE相对应的基因座,NarE是先前在脑膜炎奈瑟菌中鉴定的一种酶。由于先前描述的移码突变,淋病奈瑟菌narE的291 bp编码序列与脑膜炎奈瑟菌narE基因的部分编码序列具有100%的同一性,从而导致淋病奈瑟菌NarE蛋白N端缺失49个氨基酸。然而,我们发现了一个启动子区域和一个GTG起始密码子,通过RT-PCR和蛋白质印迹分析表明该蛋白能够表达。使用淋病奈瑟菌NarE-6xHis融合蛋白,我们证明淋病奈瑟菌酶可进行自身ADP核糖基化,但程度低于脑膜炎奈瑟菌NarE。我们还观察到,以胍丁胺和无细胞宿主蛋白作为ADP核糖受体时,淋病奈瑟菌NarE表现出ADP核糖基转移酶活性,但其活性受到人β-防御素的抑制。综上所述,我们的结果表明淋病奈瑟菌的NarE是一种功能性酶,具有细菌ADP核糖基化酶的关键特征。