Inactivation of alpha 2-antiplasmin by limited reaction with cis-dichlorodiammineplatinum (II)
alpha 2-Antiplasmin (alpha 2AP), a serpin proteinase inhibitor with two methionine residues in its reactive center, was treated with cis-dichlorodiammineplatinum (II) (cis-DDP). This compound has been utilized previously to specifically modify methionine residues. After reaction, the alpha 2AP demonstrated decreased inhibitory activity against plasmin, miniplasmin, trypsin and chymotrypsin. The reduction in activity depended on the concentration of cis-DDP; however, the amount of activity retained by the treated alpha 2AP was equivalent with each of the four proteinases. alpha 2AP that was incubated with 1.0 mM cis-DDP for 3 h at 37 degrees C was 90% inactivated. These same conditions resulted in the binding of only 1.0-1.5 mol of platinum per mol of inhibitor. In experiments with lower concentrations of cis-DDP, the amount of incorporated platinum directly correlated with the amount of inactivated alpha 2AP (1:1 stoichiometry). Reactions and functions of alpha 2AP that do not result in proteinase inhibition were not affected by cis-DDP. Cleavage of alpha 2AP by elastase, which occurs near the proteinase inhibition site, was unaffected. In addition, the affinity of alpha 2AP for the K1-3 region of plasminogen remained unchanged after treatment. These data strongly suggest that the reaction of alpha 2AP with cis-DDP involves principally a single site on the inhibitor and that this site is critical for proteinase inhibitory activity. The most likely candidate is the P'1 methionine which is adjacent to the peptide bonds cleaved in the proteinase inhibitory reactions but not in the elastase reaction.
Geary
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WA
Department of Pathology, University of Virginia Medical Center, Charlottesville 22908.
Gonias
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SL
D001665:Binding Sites / Q000187:drug effects; D002918:Chymotrypsin / Q000037:antagonists & inhibitors; D002945:Cisplatin / Q000494:pharmacology*; D004591:Electrophoresis, Polyacrylamide Gel; D005341:Fibrinolysin / Q000037:antagonists & inhibitors / Q000378:metabolism; D006801:Humans; D008715:Methionine; D010196:Pancreatic Elastase / Q000378:metabolism; D010446:Peptide Fragments / Q000037:antagonists & inhibitors; D010958:Plasminogen / Q000378:metabolism; D010984:Platinum / Q000378:metabolism; D014361:Trypsin Inhibitors; D000979:alpha-2-Antiplasmin / Q000378:metabolism* / Q000494:pharmacology
D010446:Peptide Fragments; D014361:Trypsin Inhibitors; D000979:alpha-2-Antiplasmin; C019493:miniplasmin; D010984:Platinum; D010958:Plasminogen; D008715:Methionine; D002918:Chymotrypsin; D010196:Pancreatic Elastase; D005341:Fibrinolysin; D002945:Cisplatin