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Interaction of heparin with myosin ATPase: possible involvement with the hemorrhagic activity and a correlation with
I L Tersariol1, C P Dietrich, H B Nader
1Departamento de Bioquímica, Escola Paulista de Medicina, São Paulo, Brazil.
Abstract:
Up to 50% of [35S]-heparin molecules prepared from rat skin bind to rabbit muscle myosin ATPase, in a concentration dependent manner, producing a stable complex with a dissociation constant of 3 x 10(-7) M. The [35S]-heparin in the complex has a distinct electrophoretic behaviour and is precipitated by TCA together with myosin. Other [35S]-glycosaminoglycans, namely, heparan sulfate, dermatan sulfate and chondroitin sulfate also prepared from rat tissues are unable to form complexes with the enzyme. Among the sulfated glycosaminoglycans obtained from different sources only heparin is able to displace the bound [35S]-heparin from the ATPase. Heparin with high affinity for antithrombin III, prepared by antithrombin-affinity chromatography, dislodges up to 90% of the bound [35S]-heparin. Furthermore, antithrombin III-high affinity heparin shows a high affinity for myosin ATPase when compared to antithrombin III-low affinity heparin which shows a low affinity for the enzyme. It is also shown that myosin ATPase inhibits the "in vitro" plasma anticoagulant activity of heparin. These are suggestive that the special structure of the heparin molecules needed for the binding to antithrombin and myosin ATPase bears important similarities. The mechanism of the hemorrhagic effect of heparin is discussed in view of these interactions.
Insights
Heparin binds to myosin ATPase, forming a stable complex. This interaction suggests structural similarities between heparin binding sites for antithrombin and myosin ATPase, potentially explaining heparin
Area of Science:
- Biochemistry
- Molecular Biology
- Pharmacology
Background:
- Heparin is a widely used anticoagulant.
- Its interaction with various proteins is crucial for its biological activity.
- Myosin ATPase is an enzyme involved in muscle contraction.
Purpose of the Study:
- To investigate the binding of heparin to rabbit muscle myosin ATPase.
- To characterize the nature of the heparin-myosin ATPase complex.
- To explore the structural basis for heparin's interaction with myosin ATPase and antithrombin III.
Main Methods:
- Preparation of [35S]-heparin from rat skin.
- Binding assays to determine complex formation and dissociation constant.
- Electrophoresis and precipitation techniques to analyze the complex.
- Competition assays using other glycosaminoglycans and heparin fractions.
Main Results:
- Up to 50% of [35S]-heparin bound to myosin ATPase in a concentration-dependent manner, forming a stable complex (Kd = 3 x 10(-7) M).
- Only heparin, not other glycosaminoglycans, formed complexes with myosin ATPase.
- High-affinity heparin for antithrombin III showed stronger binding to myosin ATPase and could displace more bound [35S]-heparin.
- Myosin ATPase inhibited the in vitro anticoagulant activity of heparin.
Conclusions:
- Heparin specifically binds to rabbit muscle myosin ATPase.
- The structural requirements for heparin binding to myosin ATPase are similar to those for antithrombin III.
- These interactions may contribute to the understanding of heparin's mechanism of action, including its hemorrhagic effects.