Yamada Takayuki, Fujimori Yuta, Suzuki Atsuo, Miyawaki Yuhri, Takagi Akira, Murate Takashi, Sano Masayuki, Matsushita Tadashi, Saito Hidehiko, Kojima Tetsuhito
Department of Pathophysiological Laboratory Sciences, Nagoya University Graduate School of Medicine, Nagoya, Japan.
Am J Hematol. 2009 Nov;84(11):738-42. doi: 10.1002/ajh.21532.
Combined deficiency of coagulation factor V (FV) and factor VIII (FVIII) (F5F8D) is an inherited bleeding disorder characterized by a reduction in plasma concentrations of FV and FVIII. F5F8D is genetically linked to mutations in either LMAN1 or MCFD2. Here, we investigated the molecular basis of F5F8D in a Japanese patient, and identified a novel missense mutation (p.Trp67Ser, c.200G>C) in the LMAN1, but no mutation in the MCFD2. The amount of LMAN1 in Epstein-Barr virus-immortalized lymphoblasts from the patient was found to be almost the same as that in cells from a normal individual. Interestingly, an anti-MCFD2 antibody did not co-immunoprecipitate the mutant LMAN1 with MCFD2 in lymphoblasts from the patient, suggesting the affinity of MCFD2 for the mutant LMAN1 is weak or abolished by the binding of the anti-MCFD2 antibody. In addition, a Myc/6xHis-tagged recombinant form of wild-type LMAN1 could bind to D-mannose, but that of the mutant could not. The p.Trp67Ser mutation was located in the carbohydrate recognition domain (CRD), which is thought to participate in the selective binding of LMAN1 to the D-mannose of glycoproteins as well as the EF-motif of MCFD2. Taken together, it was suggested that the p.Trp67Ser mutation might affect the molecular chaperone function of LMAN1, impairing affinity for D-mannose as well as for MCFD2, which may be responsible for F5F8D in the patient. This is the first report of F5F8D caused by a qualitative defect of LMAN1 due to a missense mutation in LMAN1. Am. J. Hematol. 2009. (c) 2009 Wiley-Liss, Inc.
凝血因子V（FV）和凝血因子VIII（FVIII）联合缺乏症（F5F8D）是一种遗传性出血性疾病，其特征是血浆中FV和FVIII浓度降低。F5F8D在基因上与LMAN1或MCFD2的突变相关。在此，我们研究了一名日本患者F5F8D的分子基础，在LMAN1中鉴定出一个新的错义突变（p.Trp67Ser，c.200G>C），但在MCFD2中未发现突变。发现该患者的爱泼斯坦-巴尔病毒永生化淋巴细胞中LMAN1的量与正常个体细胞中的量几乎相同。有趣的是，抗MCFD2抗体在该患者的淋巴细胞中不能将突变型LMAN1与MCFD2共免疫沉淀，这表明MCFD2与突变型LMAN1的亲和力较弱或因抗MCFD2抗体的结合而丧失。此外，Myc/6xHis标签的野生型LMAN1重组形式可以与D-甘露糖结合，但突变型则不能。p.Trp67Ser突变位于碳水化合物识别结构域（CRD），该结构域被认为参与LMAN1与糖蛋白的D-甘露糖以及MCFD2的EF基序的选择性结合。综上所述，提示p.Trp67Ser突变可能影响LMAN1的分子伴侣功能，损害其对D-甘露糖以及MCFD2的亲和力，这可能是该患者F5F8D的原因。这是关于由LMAN1错义突变导致的LMAN1定性缺陷引起的F5F8D的首次报道。《美国血液学杂志》2009年。（c）2009威利-利斯公司。
