Stamnaes J, Fleckenstein B, Lund-Johansen F, Sollid L M
Centre for Immune Regulation, Institute of Immunology, University of Oslo and Rikshospitalet University Hospital, Oslo, Norway.
Scand J Immunol. 2008 Nov;68(5):534-42. doi: 10.1111/j.1365-3083.2008.02173.x. Epub 2008 Sep 18.
The multifunctional enzyme transglutaminase 2 (TG2) can be located intracellularly, in the extracellular matrix (ECM) and on the cell surface. Cell surface TG2 (csTG2) is poorly recognized both by most TG2-specific commercial antibodies and celiac disease-associated anti-TG2 autoantibodies. The recent characterization of a csTG2-specific monoclonal antibody (mAb), which did not recognize ECM-associated TG2, suggested major conformational differences between csTG2 and TG2 found in the ECM. Subsequent findings based on this antibody indicated ubiquitous abundance and novel roles of csTG2 in innate immune responses. We wished to identify the epitope of 6B9 so as to shed light on the disparate antibody binding properties of csTG2- and ECM-associated TG2. Surprisingly, and despite thorough effort, we were unable to isolate TG2 as the antigen of 6B9. We found that 6B9 does not react with recombinant human TG2. In immunoprecipitation experiments, 6B9 pulled down an 85 kDa protein which was identified as CD44 by mass spectrometry. Several flow cytometry experiments including the testing of CD44s transfectants indicated that CD44, and not csTG2, is the antigen of 6B9. We conclude that 6B9 does not recognize csTG2 but rather the cell surface glycoprotein CD44. Thus, recent knowledge of csTG2 gained through the use of 6B9 should be reevaluated.
多功能酶转谷氨酰胺酶2（TG2）可定位于细胞内、细胞外基质（ECM）和细胞表面。大多数TG2特异性商业抗体和与乳糜泻相关的抗TG2自身抗体对细胞表面TG2（csTG2）的识别效果都很差。最近鉴定出一种csTG2特异性单克隆抗体（mAb），它不识别与ECM相关的TG2，这表明csTG2与在ECM中发现的TG2之间存在主要的构象差异。基于该抗体的后续研究结果表明csTG2在天然免疫反应中普遍存在且具有新的作用。我们希望鉴定出6B9的表位，以阐明csTG2和与ECM相关的TG2不同的抗体结合特性。令人惊讶的是，尽管我们付出了巨大努力，但仍无法分离出TG2作为6B9的抗原。我们发现6B9不与重组人TG2反应。在免疫沉淀实验中，6B9沉淀出一种85 kDa的蛋白质，通过质谱鉴定为CD44。包括对CD44s转染子进行检测在内的多个流式细胞术实验表明，6B9的抗原是CD44，而非csTG2。我们得出结论，6B9不识别csTG2，而是识别细胞表面糖蛋白CD44。因此，通过使用6B9获得的关于csTG2的最新认识应重新评估。
