Extended interactions with prothrombinase enforce affinity and specificity for its macromolecular substrate.
J Biol Chem
; 277(48): 46191-6, 2002 Nov 29.
Article
de En
| MEDLINE
| ID: mdl-12370181
ABSTRACT
The specific action of serine proteinases on protein substrates is a hallmark of blood coagulation and numerous other physiological processes. Enzymic recognition of substrate sequences preceding the scissile bond is considered to contribute dominantly to specificity and function. We have investigated the contribution of active site docking by unique substrate residues preceding the scissile bond to the function of prothrombinase. Mutagenesis of the authentic P(1)-P(3) sequence in prethrombin 2/fragment 1.2 yielded substrate variants that could be converted to thrombin by prothrombinase. Proteolytic activation was also observed with a substrate variant containing the P(1)-P(3) sequence found in a coagulation zymogen not known to be activated by prothrombinase. Lower rates of activation of the variants derived from a decrease in maximum catalytic rate but not in substrate affinity. Replacement of the P(1) residue with Gln yielded an uncleavable derivative that retained the affinity of the wild type substrate for prothrombinase but did not engage the active site of the enzyme. Thus, active site docking of the substrate contributes to catalytic efficiency, but it is does not determine substrate affinity nor does it fully explain the specificity of prothrombinase. Therefore, extended interactions between prothrombinase and substrate regions removed from the cleavage site drive substrate affinity and enforce the substrate specificity of this enzyme complex.
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Collection:
01-internacional
Base de données:
MEDLINE
Sujet principal:
Thromboplastine
Limites:
Humans
Langue:
En
Journal:
J Biol Chem
Année:
2002
Type de document:
Article
Pays d'affiliation:
États-Unis d'Amérique