MECHANISM UNDERLYING CONGENTIAL FVII VARIANT (FVIIR79Q) WITH VARIABLE FVII : c USING TISSUE THROMBOPLASTIN FROM DIFFERENT SOURCE

为探讨FVIIR 79 Q在不同来源的组织凝血活酶作用下表现出不同的FVII：c的机制，用野生型和突变型FVII cDNA瞬时转染COS-1细胞，测定了条件培养液中FVII：c和FVII：ag的含量。pcDNAVIIR 79 W为12 ng，pcDNAVIIR 79 P为14 ng，pcDNAVIIR 79 W为40 ng。第七：用人TF和兔TF测定pcDNAVIIwt构建体转染的细胞培养液中的VII：c，结果与用人TF和兔TF测定的结果没有差异。pcDNAVIIR 79 Q和pcDNAVIIR 79 W转染细胞的VIIc为野生型的30%，而pcDNAVIIR 79 W和pcDNAVIIR 79 P转染细胞的VIIc为野生型的10%。虽然重组FVIIR 79 G的VII：c与对照组相比无明显差异，但仍有B水平 ...更多信息 用兔TF和人TF测定的重组FVIIR 79 Q、FVIIR 79 W和FVIIR 79 P的VII：c均低于人TF。人FVIIa/人TF复合物的能量最小模型结构表明，FVIIR 79 Q中的Q79与TF中的E56之间的氢键消失。TF的E24和E56与FVIIR 79 Q的Q79之间的距离估计为7 ~ 8A，推测重组FVIIR 79 Q的低VII：c是通过FVIIa/TF复合物的微观构象变化而干扰的。推测重组FVIIR 79 G、FVIIR 79 W和FVIIR 79 P的VII：c降低是由于突变型FVII在TF附近的空间限制改变。人FVIIa/兔TF复合物的建模显示类似于人FVIIa/人TF复合物。FVIIR 79 Q的能量最小化模型结构表明兔TF和人TF之间的复合物几乎没有不同。重组FVIIR 79 G的VII：c在兔TF和人TF使用之间没有差异。重组FVIIR 79 W和FVIIR 79 P与兔TF和人TF的c值存在差异，很难用能量最小化模型结构的比较来简单解释其机制。FVIIa/兔TF复合物与FVIIa/人TF复合物的差异。少

To elucidate the mechanism by which FVIIR79Q shows variable FVII : c using tissue thromboplastin from different source, transient transfections were performed in COS-1 cells with wild type and mutant FVII cDNAs and FVII : c and FVII : ag in condition media were measured.The VII : ag in the media on cells transfected with pcDNAVIIwt contained 18ng/ml, pcDNAVIIR79Q ; 22ng, pcDNAVIIR79G ; 12ng, pcDNAVIIR79W ; 14ng, pcDNAVIIR79P ; 40ng. The VII : c in the media of cells transfected with the construct of pcDNAVIIwt did not have the difference between the result by the human TF and that by the rabbit TF.However the VII : c in the media on cells transfected with variant types were lower than that of wild type when assayed both with the human TF and the rabbit TF.The VII : c in the media on cells transfected with pcDNAVIIR79Q and pcDNAVIIR79W were 30% of that of wild type, and VIIc in pcDNAVIIR79W and pcDNAVIIR79P 10%. Although the VII : c of recombinant FVIIR79G did not have diffenent level b … More etween assayed with rabbit TF and with human TF, the VII : c of recombinant FVIIR79Q, FVIIR79W and FVIIR79P assayed with rabbit TF were lower than those of human TF.The energy minimised model structure of human FVIIa/human TF complex indicates that H-bond between Q79 in FVIIR79Q and E56 in TF disappears. And the distance between E24 and E56 of TF and Q79 of FVIIR79Q is estimated to be 7 to 8A.then it is speculated that the low VII : c of recombinant FVIIR79Q is interfering by the micro conformational change of FVIIa/TF complex. It is speculated that reduced VII : c of recombinant FVIIR79G, FVIIR79W and FVIIR79P are due to change of space restriction in mutant FVII close to TF.Modeling of human FVIIa/rabbitTF complex shows similar to human FVIIa/human TF complex. The energy minimized model structure of FVIIR79Q indicates little different complex between rabbit TF and human TF.VII : c of recombinant FVIIR79G showed no different between using by rabbit TF and using by human TF.FVII : c of recombinant FVIIR79W and FVIIR79P showed different between with rabbit TF and with human TF.It was difficult simply to explain the mechanism by comparison of the energy minimized model structure of FVIIa/rabbit TF complex with that of FVIIa/human TF complex. Less