Redox modulation of integrin [correction of integin] alpha IIb beta 3 involves a novel allosteric regulation of its thiol isomerase activity.
The molecular mechanisms involved in regulating the activation-dependent conformational switch in integrins are not known although recent evidence suggests that integrins are a direct target for redox modulation. We have identified an endogenous integrin thiol isomerase activity that may be responsible for regulating integrin activation states. The purpose of this study was to examine the effects of redox conditions elicited by nitric oxide and glutathione on the thiol isomerase activity of the platelet integrin alphaIIbbeta3 and also on the activation status of this integrin in intact platelets. The universal integrin activator, Mn2+, stimulates the thiol isomerase activity in purified alphaIIbbeta3. Kinetic analysis reveals that alphaIIbbeta3 is an allosteric enzyme which displays positive cooperativity in the presence of Mn2+ with an apparent Hill coefficient of 1.9. Also, addition of Mn2+ to platelets results solely in activation of the integrin as demonstrated by the binding of the antibody PAC-1. The addition of the nitric oxide donors SNP, SIN-1, and SNOAC in combination with glutathione can directly reverse the activation state of the platelet integrin induced by Mn2+. These compounds have no effect on platelet secretory responses indicating a direct effect on the integrin. In the presence of nitric oxide and glutathione, the enzymatic activity of alphaIIbbeta3 also displays positive cooperativity (apparent Hill coefficient of 1.9), and a significant increase in the saturability of the enzyme was observed. Thus, redox agents simultaneously modulate the thiol isomerase activity of purified alphaIIbbeta3 and its active conformation in intact platelets, suggesting a molecular mechanism for integrin regulation.
Walsh
Geraldine M
GM
Department of Clinical Pharmacology, Royal College of Surgeons in Ireland, Dublin, Ireland.
Sheehan
David
D
Kinsella
Anthony
A
Moran
Niamh
N
O'Neill
Sarah
S
D000494:Allosteric Regulation / Q000187:drug effects; D001792:Blood Platelets / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism; D004789:Enzyme Activation / Q000187:drug effects; D005978:Glutathione / Q000494:pharmacology; D006801:Humans; D008345:Manganese / Q000494:pharmacology; D009569:Nitric Oxide / Q000494:pharmacology; D010084:Oxidation-Reduction; D015539:Platelet Activation / Q000187:drug effects; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000302:isolation & purification / Q000378:metabolism*; D019704:Protein Disulfide-Isomerases / Q000378:metabolism*; D012259:Ribonuclease, Pancreatic / Q000378:metabolism; D013917:Thrombin / Q000494:pharmacology
D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D009569:Nitric Oxide; D008345:Manganese; D012259:Ribonuclease, Pancreatic; D013917:Thrombin; D019704:Protein Disulfide-Isomerases; D005978:Glutathione