Sislic acid recognition proteins of Trypanosoma species

反式唾液酸酶是一些感染性锥虫特有的酶，它催化唾液酸从宿主衍生的糖结合物转移到寄生虫表面粘蛋白样受体分子，反之亦然。一些证据表明，寄生虫和宿主细胞表面的反式唾液酸酶和唾液酸化分子对于寄生虫入侵宿主细胞和逃离宿主防御系统是重要的。我们从分子水平上对反式唾液酸酶进行了功能分析。在本研究中，我们成功地从引起非洲昏睡病的布氏锥虫的培养液中纯化并鉴定了反式唾液酸酶。这种酶与查加斯病的病原体克氏锥虫有几个生化上的相似之处。我们还利用细菌表达系统对克鲁兹毛滴虫重组反式唾液酸酶进行了均相表达和纯化。这些酶用于宿主细胞对寄生虫蛋白的识别研究。比较纯化的重组蛋白的活性和失活形式对于了解反式唾液酸酶活性的功能具有重要意义。我们获得并分析了几个在该寄生虫的锥虫鞭毛期和上鞭毛期表达的克氏锥虫反式唾液酸酶基因。在感染性类鞭毛虫期(T-TS)表达的酶通过糖基磷脂酰肌醇(GPI)锚点定位在寄生虫表面，在体外很容易进入培养上清，在体内很容易进入动物的血液。另一方面，非感染性上胚体阶段的反式唾液酸酶不会释放到介质中，也不会被抗T-TS羧基末端重复序列的抗体识别。我们对这些家族基因进行了测序，以了解这两种不同类型的反式唾液酸酶的功能、表达和定位。

The unique enzyme of some infective Trypanosoma species, trans-sialidase catalyzes the sialic acid transfer reactions from host derived glycoconjugates to the parasite surface mucinlike acceptor molecules and vice versa. Several lines of evidence have suggested that trans-sialidase and sialylated molecules on the parasite and host cell surface are important for parasite invasion to the host cells and escape from host defense systems. We have investigated the functional analysis of trans-sialidase in molecular level.In this study, we have successfully purified and characterized trans-sialidase obtained from the culture medium of Trypanosome brucei, which causes African sleeping sickness. This enzyme showed several biochemical similarities to that of Trypanosoma cruzi, which is the etiological agent of Chagas' disease. We also have expressed and purified homogeneously the recombinant trans-sialidase of T. cruzi using bacterial expression system. These enzymes are used for the recognition studies of the parasite protein by host cells. The comparison of enzymatically active and inactive forms of purified recombinant proteins were important for understanding the function of trans-sialidase activitis.We have obtained and analyzed several T. cruzi trans-sialidase genes expressed in trypomastigote and epimastigote stages of this parasite. The enzyme expressed in infective trypomastigote stage (T-TS) is located on the parasite surface through a glycosyl phosphatididylinositol (GPI) anchor, and readily shed into culture supernatant in vitro and into the bloodstream of animals in vivo. On the other hands, trans-sialidase of noninfective epimastigote stage is not released into the medium nor recognized by antibodies against the carboxyl terminal repeats of T-TS.We have sequences these family genes to understand functions, molecular mechanisms of expression and localization of these two different types of trans-sialidases.