The gelation kinetics of platelet extracts |
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Authors: | Chauying J. Jen Larry V. McIntire |
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Affiliation: | Biomedical Engineering Laboratory, Chemical Engineering Department, Rice University, P.O. Box 1892, Houston, TX 77251 U.S.A. |
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Abstract: | The kinetics of the gelation process that occurs upon warming cold platelet extracts were studied using a sensitive rheometer. At micromolar or less free Ca2+ concentrations and in the presence of 1 mM ATP, the gel rigidity curves showed several peaks, indicating that platelet extract proteins went through network assembling/disassembling cycles during gelation. The gelation kinetics were accelerated by increasing the free Ca2+ concentration up to about 2 μM. At 4–15 μM free Ca2+, the gelation cycles were completely abolished except for the first peak. The gelation process became one of monotonically increasing elastic modulus at millimolar free Ca2+ concentrations. Trifluoperazine (50 μM), a calmodulin inhibitor, did not affect gelation at micromolar free Ca2+ concentrations. Except for the first gelation step, which was completed within 5 min after warming, the rest of the gelation process was found to be affected by K+, ATP, cytochalasin E and colchicine. K+ at concentrations higher than 10 mM retarded the gelation kinetics. Extracts prepared with low (0.1 mM) ATP content showed impaired gelations, and this was partially reversed by adding 1 mM ATP, but not 1 mM adenylylimidodiphosphate (p[NH]ppA). Both cytochalasin E (1 μM) and colchicine (1 mM) interfered with the gelation process. |
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Keywords: | Gelation kinetics Protein network Rheology Viscoelasticity (Platelet extract) p[NH]ppA adenylylimidodiphosphate EGTA Pipes 1,4-piperazinediethanesulfonic acid SDS sodium dodecyl sulfate |
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