Important role of arginine 129 in heparin-binding site of antithrombin III. Identification of a novel mutation arginine 129 to glutamine.
An hereditary abnormal antithrombin III (ATIII Geneva) with defective heparin cofactor activity was characterized by DNA single strand amplification and subsequent direct sequencing. ATIII Geneva was found to have a G to A transition in Exon IIIa leading to an Arg-129 to Gln mutation. This amino acid is part of the ATIII region comprising residues 114-154, which contains the highest proportion of basic residues (Arg or Lys), and is known from chemical modification studies to be involved in heparin binding. The variant protein did not bind heparin-Sepharose and was isolated from the propositus plasma by immunoaffinity chromatography. High affinity (for ATIII) heparin had only a minimal effect on thrombin and activated factor X inhibition by the purified abnormal ATIII. Taken together, these results demonstrate an important role for Arg-129 in the binding and interaction of ATIII with heparin of high affinity. We propose that a cooperation between Lys-125, Arg-129, Lys-136, and Arg-47 exposed at the surface of the inhibitor allows the binding of the essential pentasaccharide domain of heparin which is specific for the ATIII interaction.
Gandrille
S
S
Laboratoire d'Hématologie, Faculté des Sciences Pharmaceutiques et Biologiques, Paris, France.
Aiach
M
M
Lane
D A
DA
Vidaud
D
D
Molho-Sabatier
P
P
Caso
R
R
de Moerloose
P
P
Fiessinger
J N
JN
Clauser
E
E
D000595:Amino Acid Sequence; D000990:Antithrombin III / Q000235:genetics* / Q000378:metabolism; D001120:Arginine; D001483:Base Sequence; D001665:Binding Sites; D004247:DNA / Q000097:blood / Q000235:genetics / Q000302:isolation & purification; D005091:Exons; D005973:Glutamine; D006493:Heparin / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D007962:Leukocytes / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009154:Mutation; D016133:Polymerase Chain Reaction; D011487:Protein Conformation
D005973:Glutamine; D000990:Antithrombin III; D006493:Heparin; D004247:DNA; D001120:Arginine