Study on complex formation between recombinant human thrombomodulin fragment and thrombin using surface plasmon resonance.
Human thrombomodulin (hTM) is a newly described endothelial cell associated protein that functions as a potent natural anticoagulant by converting thrombin from a procoagulant protease to an anticoagulant. The affinity constant of recombinant human soluble TM (rhs-TM) and a peptide containing active site of hTM fragment (f-hTM) with thrombin were determined using the surface plasmon resonance. The interaction of f-hTM with thrombin could be analyzed by a simple model, whereas the association and the dissociation steps of rhs-TM with thrombin consisted of at least two kinds of interaction phases. The dissociation constant for complex (K(D)) of f-hTM and thrombin was determined to be 205 nM, which was more than twice as high as that of rhs-TM (6.7 and 75 nM). The lower affinity of f-hTM was not due to the slow association rate but to the rapid dissociation rate. It comes clear that f-hTM interacts with thrombin rapidly.
Kishida
A
A
Department of Applied Chemistry and Chemical Engineering, Faculty of Engineering, Kagoshima University, Korimoto, Kagoshima, Japan.
Nakashima
M
M
Sakamoto
N
N
Serizawa
T
T
Maruyama
I
I
Akashi
M
M
D006801:Humans; D007700:Kinetics; D011994:Recombinant Proteins / Q000032:analysis; D020349:Surface Plasmon Resonance; D013917:Thrombin / Q000032:analysis*; D018180:Thrombomodulin / Q000032:analysis*
D011994:Recombinant Proteins; D018180:Thrombomodulin; D013917:Thrombin