Role of the gamma-carboxyglutamic acid domain of activated factor X in the presence of calcium during inhibition by antithrombin-heparin.
BACKGROUND
Factor (F)Xa has 11 gamma-carboxylated glutamic acid (Gla) residues that are involved in calcium-dependent membrane binding. The serpin antithrombin (AT) is an important physiological regulator of FXa activity in an inhibition reaction that is enhanced by heparin. Recently, Rezaie showed that calcium further enhanced the heparin-catalyzed AT inhibition of FXa by promoting 'ternary complex' formation, and these results showed a role for the gamma-carboxyl-glutamate (Gla)-domain of FXa.


OBJECTIVE
In this study, we used recombinant FXa mutants to assess the role of individual Gla residues in augmenting or antagonizing the AT-heparin inhibition reaction in the presence of calcium.


CONCLUSIONS
In the absence of heparin, AT inhibition of plasma and the recombinant FXas were essentially equivalent. Similar to plasma-derived FXa, calcium increased about 3-fold the inhibition rate of wild-type recombinant FXa by AT-heparin over that in the presence of EDTA. Interestingly, three different effects were found with the recombinant FXa Gla-mutants for AT-heparin inhibition: (i) Gla-->Asp 14 and 29 were enhanced without calcium; (ii) Gla-->Asp 16 and 26 were not enhanced by calcium; and (iii) Gla-->Asp 19 was essentially the same as wild-type recombinant FXa. These results support a theory that mutating individual Gla residues in FXa alters the calcium-induced conformational changes in the Gla region and affects the antithrombin-heparin inhibition reaction.
Whinna
H C
HC
Department of Pathology and Laboratory Medicine, Carolina Cardiovascular Biology Center, The University of North Carolina School of Medicine, Chapel Hill, NC 27599, USA. whinna@med.unc.edu
Lesesky
E B
EB
Monroe
D M
DM
High
K A
KA
Larson
P J
PJ
Church
F C
FC
D015055:1-Carboxyglutamic Acid / Q000502:physiology*; D019943:Amino Acid Substitution; D000991:Antithrombins / Q000494:pharmacology*; D001665:Binding Sites; D002118:Calcium / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D015951:Factor Xa / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D006493:Heparin / Q000378:metabolism / Q000494:pharmacology*; D006801:Humans; D008958:Models, Molecular; D011485:Protein Binding; D017434:Protein Structure, Tertiary / Q000502:physiology
D000991:Antithrombins; D015055:1-Carboxyglutamic Acid; D006493:Heparin; D015951:Factor Xa; D002118:Calcium