Bramley J, Demarco de Hormaeche R, Constantinidou C, Nassif X, Parsons N, Jones P, Smith H, Cole J
School of Biochemistry, University of Birmingham, U.K.
Microb Pathog. 1995 Mar;18(3):187-95. doi: 10.1016/s0882-4010(95)90040-3.
A stable, sialyltransferase-deficient mutant of Neisseria gonorrhoeae strain F62 totally defective in CMP-NANA-dependent lipopolysaccharide (LPS) sialylation was isolated by insertion mutagenesis with transposon Tn 1545-delta 3 and screened for unlabelled colonies following incubation with CMP-14C-NANA. In contrast to the parental strain which became serum resistant on incubation with CMP-NANA or blood cell extracts, the mutant, JB1, remained serum sensitive. French press extracts of strain F62 catalysed LPS sialylation, but corresponding extracts of the mutant were inactive. Five LPS components were detected by SDS-PAGE in the parental strain. Five components of the same Mr were also found in the mutant. Three identical components were detected by Western blotting using MAb 3F11, which recognizes the Gal beta 1-4GlcNAc groups in the conserved LPS components of F62 which can be sialylated. The mutant, JB1, is therefore deficient in the sialyltransferase that is essential for both LPS sialylation and conversion of serum-sensitive gonococci to serum resistance by either CMP-NANA or blood cell extracts. No evidence was obtained for an LPS sialylation pathway by blood cell extracts that is independent of CMP-NANA.
通过用转座子Tn 1545-δ3进行插入诱变，分离出淋病奈瑟菌F62株的一种稳定的、唾液酸转移酶缺陷型突变体，该突变体在CMP-N-乙酰神经氨酸（CMP-NANA）依赖性脂多糖（LPS）唾液酸化方面完全缺陷，并在与CMP-14C-NANA孵育后筛选未标记的菌落。与用CMP-NANA或血细胞提取物孵育后产生血清抗性的亲本菌株不同，突变体JB1仍然对血清敏感。F62株的法国压榨机提取物催化LPS唾液酸化，但突变体的相应提取物无活性。通过SDS-PAGE在亲本菌株中检测到五种LPS成分。在突变体中也发现了相同分子量的五种成分。使用单克隆抗体3F11通过蛋白质印迹检测到三种相同的成分，该抗体识别F62保守LPS成分中可被唾液酸化的Galβ1-4GlcNAc基团。因此，突变体JB1缺乏唾液酸转移酶，该酶对于LPS唾液酸化以及通过CMP-NANA或血细胞提取物将血清敏感的淋球菌转化为血清抗性都是必不可少的。没有获得血细胞提取物存在独立于CMP-NANA的LPS唾液酸化途径的证据。
