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

2000 Fiscal Year Final Research Report Summary

• Project/Area Number: 11672295
• Research Category: Grant-in-Aid for Scientific Research (C)
• Allocation Type: Single-year Grants
• Section: 一般
• Research Field: Laboratory medicine
• Research Institution: SHINSHU UNIV., SCHOOL OF ALLIED MEDICAL SCIENCES
• Principal Investigator: TAKAMIYA Osamu SHINSHU UNIV., SCHOOL OF ALLIED MEDICAL SCIENCES PROF., 医療技術短期大学部, 教授 (50216785)
• Project Period (FY): 1999 – 2000
• Keywords: HUMAN COAGULATION FACTOR VII / FACTOR VII COAULATION ACTIVITY / TISSUE FACTOR

Research Abstract

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 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.