Study on the mode of carbohydrate recognition by functional analysis of mannan-binding protein as a multi-functional protein.

甘露聚糖结合蛋白(MBP)是一种C型血清凝集素，专用于甘露糖、岩藻糖和N-乙酰氨基葡萄糖，与清除宿主动物中的病原微生物有关。虽然猪被认为是一种有吸引力的异种可移植组织和器官来源，但人们对猪的MBP知之甚少。在我们以前的研究中，发现磷甘露聚糖而不是甘露聚糖能有效地抑制新生仔猪血清对一些粗制革兰氏阴性菌的C1q非依赖性杀菌活性。在此，我们从新生和成年猪血清中分离并鉴定了一种新的33 kDa还原条件下的磷甘露聚糖结合凝集素(PMBL)。猪PMBL通过与甘露聚糖和甘露聚糖结合而触发的凝集素途径激活补体系统，而与MBP不同的是，含有6-磷酸甘露糖的寡糖有效地抑制了补体系统的激活。我们的观察结果表明，猪Pmbl在…中起重要作用从新生儿到成年，CAL在先天免疫防御中的作用更大。MBP主要在肝脏合成，天然存在两种形式：血清MBP(S-MBP)和细胞内MBP(I-MBP)。S-丝氨酸蛋白通过凝集素途径激活补体，并与丝氨酸蛋白酶结合。另一方面，I-MBP的亚细胞定位及其功能意义尚不清楚。在这里，我们证明了I-MBP在细胞质颗粒中有明显的积聚，并且主要定位于内质网，并参与COPII囊泡介导的内质网到高尔基体的转运。然而，无论是缺乏糖结合活性的突变体(C236S/C244S)I-MBP还是野生型I-MBP在衣霉素处理的细胞中的亚细胞定位都显示出同样弥散的细胞质分布，这表明I-MBP在ER和COPII囊泡中的独特积累是由N-糖-凝集素相互作用介导的。这些结果有力地表明，I-MBP在糖蛋白质量控制中可能作为一种货物运输凝集素，促进内质网到高尔基体的运输。此外，我们正在制备识别表达在人结直肠癌SW1116细胞表面的MBP配体寡糖的单抗。较少

Mannan-binding protein (MBP) is a C-type serum lectin specific for mannose, fucose and N-acetylglucosamine, and is associated with the removal of pathogenic microorganisms from the host animals. While the pig has been proposed to be an attractive source of xeno transplantable tissues and organs, little is known about porcine MBP. In our previous studies, phosphomannan, but not mannan, was found to be an effective inhibitor of the C1q-independent bactericidal activity of newborn piglet serum against some rough strains of gram-negative bacteria. Here we show that a novel phosphomannan-binding lectin (PMBL) of 33 kDa under reducing conditions was isolated from both newborn and adult porcine serum and characterized. Porcine PMBL activated the complement system via the lectin pathway triggered by binding with both phosphomannan and mannan, which, unlike MBP, was effectively inhibited by mannose 6-phosphate-containing oligosaccharides. Our observations suggest that porcine PMBL plays a criti … More cal role in the innate immune defense from the newborn stage to adulthood. MBP is mainly synthesized in the liver and occurs naturally in two forms, serum MBP (S-MBP) and intracellular MBP (I-MBP). S-MBP activates complement in association with MBP-associated serine proteases via the lectin pathway. On the other hand, the subcellular localization of I-MBP and its functional implication have not been clarified yet. Here, we demonstrated that I-MBP shows distinct accumulation in cytoplasmic granules, and is predominantly localized in the ER and involved in COPII vesicle-mediated ER-to-Golgi transport. However, the subcellular localization of either a mutant (C236S/C244S) I-MBP, which lacks carbohydrate-binding activity, or the wild-type I-MBP in tunicamycin-treated cells shows an equally diffuse cytoplasmic distribution, suggesting that the unique accumulation of I-MBP in the ER and COPII vesicles is mediated by an N-glycan-lectin interaction. These results strongly indicate that I-MBP may function as a cargo transport lectin facilitating ER-to-Golgi traffic in glycoprotein quality control. In addition, we are preparing monoclonal antibodies recognizing MBP-ligand oligosaccharides expressed on the surfaces of a human colorectal carcinoma SW1116 cells. Less