Lee Hyun-Ku, Lee Jongdae, Tobias Peter S
Department of Immunology, The Scripps Research Institute, La Jolla, CA 92037, USA.
J Immunol. 2002 Apr 15;168(8):4012-7. doi: 10.4049/jimmunol.168.8.4012.
Toll-like receptor 2 (TLR2)-mediated cell activation induced by commercial preparations of LPS was recently shown to arise from impurities whose identities are not known. In this work, we determined the molecules responsible for TLR2-mediated cell activation in LPS derived from Escherichia coli K-12 strain LCD25. When LCD25 LPS was phenol extracted, two proteins capable of TLR2-mediated cell activation were purified and identified as E. coli lipoproteins. We cloned, expressed, and purified these two lipoproteins, Lip19 and Lip12. Lip19 or Lip12 activated TNF-alpha production from RAW264.7 and THP-1 cells in a TLR2-dependent manner. However, neither Lip19 nor Lip12 activated HUVECs, which lack endogenous TLR2. Additionally, IkappaB kinase beta and c-Jun N-terminal kinase 1 activation in THP-1 cells induced by Lip19 or Lip12 was observed. TLR2 activation by Lip19 and Lip12 in HEK293 cells was blocked by inhibitory anti-TLR2 mAbs. The unlipidated mutants, Lip19-C19S and Lip12-C21S, in which the NH(2)-terminal cysteine was substituted by serine, lost their ability to activate TLR2-transfected HEK 293 cells. Taken together, these results demonstrate that two lipoproteins constitute the major contaminants responsible for TLR2-mediated cell activation in E. coli LCD25 LPS.
最近研究表明，商业制备的脂多糖（LPS）所诱导的Toll样受体2（TLR2）介导的细胞活化是由身份不明的杂质引起的。在本研究中，我们确定了源自大肠杆菌K-12菌株LCD25的LPS中负责TLR2介导细胞活化的分子。对LCD25 LPS进行酚提取后，纯化出两种能够介导TLR2细胞活化的蛋白质，并鉴定为大肠杆菌脂蛋白。我们克隆、表达并纯化了这两种脂蛋白，即Lip19和Lip12。Lip19或Lip12以TLR2依赖的方式激活RAW264.7和THP-1细胞产生肿瘤坏死因子-α（TNF-α）。然而，Lip19和Lip12均未激活缺乏内源性TLR2的人脐静脉内皮细胞（HUVECs）。此外，观察到Lip19或Lip12诱导THP-1细胞中的IκB激酶β和c-Jun氨基末端激酶1活化。在HEK293细胞中，Lip19和Lip12对TLR2的激活被抑制性抗TLR2单克隆抗体阻断。未脂化的突变体Lip19-C19S和Lip12-C21S，其中氨基末端半胱氨酸被丝氨酸取代，失去了激活TLR2转染的HEK 293细胞的能力。综上所述，这些结果表明两种脂蛋白是大肠杆菌LCD25 LPS中负责TLR2介导细胞活化的主要污染物。
