Adhesin of Helicobacter pylon : Identification and therapeutic application

幽门螺杆菌（Helicobacter pylon，HP）在胃粘膜上的附着是HP感染持续存在的关键步骤.生物体与上皮的紧密粘附深刻地影响上皮功能，引起各种疾病。这种附着是由一种称为粘附素的特殊细菌蛋白介导的，它能识别特定的受体。在大多数情况下，细胞表面糖缀合物已被证明是细菌粘附受体。在HP的情况下，已经报道了几种具有不同受体特异性的假定粘附素。在这个项目中，我们试图确定粘附素特异性识别硫苷脂。采用两种不同的层析方法，一种是肝素-琼脂糖凝胶层析，另一种是硫酸纤维素层析，从HP的超声处理上清液中鉴定了两种候选蛋白。通过对HP基因组中同源序列的搜索，获得了HP亲和纯化蛋白的部分氨基酸序列，其中一个被鉴定为热休克蛋白60（HSP），另一个被鉴定为功能未知的蛋白。由于其他研究者假设HSP作为一种假定的粘附素作用于细菌粘附的初始步骤，我们克隆了HSP基因以阐明HSP的结合特异性。这项工作正在进行中。其次，我们试图表征蒙古沙鼠的粘附受体，蒙古沙鼠被认为是HP感染诱导的人类胃溃疡和癌症的合适动物模型。在该动物中，与小鼠相比，硫苷脂在胃粘膜中大量表达，表明在该动物中观察到的较高数量的细菌附着可能是由于受体丰度。现将这一结果提交出版。为了进一步证实硫苷脂作为受体的作用，我们克隆了人磺基转移酶。在硫酸脂缺乏的细胞中表达的磺基转移酶现在正在进行中。

Attachment of Helicobacter pylon (HP) to gastric epithelium is a crucial step for perpetuating the infection. Tight adhesion of the organism to the epithelium profoundly affects the epithelial function causing various diseases. The attachment is mediated by specific bacterial proteins called adhesins which recognize specific receptors. In most cases, cell surface glycoconjugates have been demonstrated as bacterial adhesion receptors. In the case of HP, several putative adhesins with different receptor specificities have been reported. In this project, we tried to identify adhesins which specifically recognize sulfatide. Using two different affimty chromatographies, one with heparin-sepharose and the other with sulfated cellulose, two candidate proteins from sonicated supernatant from HP were identified. Several partial amino acid sequences of the peptides derived from affinity-purified proteins were obtained By searching homologous sequences in the genome data of HP, one was identified as heat shock protein 60 (HSP) and the other as a functionally unknown protein. Since other researchers hypothesized HSP as a putative adhesin acting at the initial step of bacterial attachment, we cloned HSP gene to clarify the binding specificity of HSP. This work is now in progress. Secondary, we tried to characterize the adhesion receptors in Mongolian gerbils which have been known as a suitable animal model of human gastric ulcer and cancer induced by HP infection. In this animal, sulfatides were abundantly expressed in the gastric mucosa compared with mice, indicating the higher number of bacterial attachment seen in this animal may be due to receptor abundance. This result is now submitted for publication. To further confirm the role of sulfatide as a receptor, we have cloned human sulfotransferase. The expression of sulfotransferase in the sulfatide-deficient cells is now in progress.