TY - JOUR
T1 - Enhancing the activity of protein C by mutagenesis to improve the membrane-binding site
T2 - Studies related to proline-10
AU - Shen, L.
AU - Shah, A. M.
AU - Dahlback, B.
AU - Nelsestuen, G. L.
N1 - Copyright:
Copyright 2007 Elsevier B.V., All rights reserved.
PY - 1997
Y1 - 1997
N2 - Bovine and human protein C show high homology in the amino acids of their GLA domains (amino-terminal 44 residues), despite the about 10-fold higher membrane affinity of the human protein. A proposed membrane contact site and mechanism suggested that this difference was largely due to the presence of proline at position 10 of bovine protein C versus histidine at position 10 of human protein C [McDonald, J. F., Shah, A. M., Schwalbe, R. A., Kisiel, W., Dahlback, B., and Nelsestuen, G. L. (1997) Biochemistry 36, 5120-5127]. This study examined the impact of replacing proline-10 in bovine protein C with histidine, and the reverse change in human protein C. In both cases, the protein containing proline-10 showed lower membrane affinity, about 10-fold lower for bovine protein C and 5-fold lower for human protein C. As expected, activated human protein C (hAPC) containing proline at position 10 showed 2.4-3.5-fold lower activity than wild type hAPC, depending on the assay used. Most interesting was that bovine APC containing histidine- 10 displayed up to 15-fold higher activity than wild type bAPC. This demonstrated the ability to improve both membrane contact and activity by mutation. This general strategy should be applicable to other vitamin K- dependent proteins, providing opportunities to study function as well as to produce proteins that may find use as promoters and inhibitors of blood coagulation in pathological states.
AB - Bovine and human protein C show high homology in the amino acids of their GLA domains (amino-terminal 44 residues), despite the about 10-fold higher membrane affinity of the human protein. A proposed membrane contact site and mechanism suggested that this difference was largely due to the presence of proline at position 10 of bovine protein C versus histidine at position 10 of human protein C [McDonald, J. F., Shah, A. M., Schwalbe, R. A., Kisiel, W., Dahlback, B., and Nelsestuen, G. L. (1997) Biochemistry 36, 5120-5127]. This study examined the impact of replacing proline-10 in bovine protein C with histidine, and the reverse change in human protein C. In both cases, the protein containing proline-10 showed lower membrane affinity, about 10-fold lower for bovine protein C and 5-fold lower for human protein C. As expected, activated human protein C (hAPC) containing proline at position 10 showed 2.4-3.5-fold lower activity than wild type hAPC, depending on the assay used. Most interesting was that bovine APC containing histidine- 10 displayed up to 15-fold higher activity than wild type bAPC. This demonstrated the ability to improve both membrane contact and activity by mutation. This general strategy should be applicable to other vitamin K- dependent proteins, providing opportunities to study function as well as to produce proteins that may find use as promoters and inhibitors of blood coagulation in pathological states.
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U2 - 10.1021/bi971730v
DO - 10.1021/bi971730v
M3 - Article
C2 - 9440875
AN - SCOPUS:0031424043
SN - 0006-2960
VL - 36
SP - 16025
EP - 16031
JO - Biochemistry
JF - Biochemistry
IS - 51
ER -