title: Inactivation of antiplasmin at low pH: evidence for the formation of latent molecules.
abstract: Several serine proteinase inhibitors (serpins) are metastable proteins which under certain conditions may undergo conformational changes resulting in the insertion of the reactive centre loop into the so-called Abeta-sheet and hence forming latent molecules. Here we have studied the inactivation of antiplasmin as a function of pH and temperature with time. At decreased pH (4.9-5.8) and at room temperature, antiplasmin activity decreased following first-order kinetics. Analysis by polyacrylamide gel electrophoresis under non-denaturing conditions demonstrated that only minor amounts of polymerized material formed after extensive incubation (4 days) at room temperature. However, on incubation at elevated temperatures (45 or 55 degrees C), a rapid formation of polymerized material was observed. We also demonstrated that antiplasmin inactivated by treatment at pH approximately 5 at room temperature spontaneously slowly regained some activity if incubated in a buffer of neutral pH. Furthermore, by treatment with 4 M guanidinium chloride for about 30 min, followed by dialysis against a neutral phosphate buffer, considerable activity (almost 40%) was regained. Thus, we conclude that antiplasmin, at least partially, at lower temperatures is transformed into a latent form, which could be reactivated, in a similar manner as PAI-1. At increased temperature, however, polymerization seems to be the predominant reason for inactivation.
authors.affiliation: Department of Clinical Chemistry and Blood Coagulation, Karolinska hospital, Karolinska Institute, Stockholm SE-17176, Sweden.
authors.forename: Haiyao
authors.initials: H
authors.lastname: Wang
authors.forename: Sarolta
authors.initials: S
authors.lastname: Pap
authors.forename: Björn
authors.initials: B
authors.lastname: Wiman
mesh_terms: D019281:Dimerization; D019791:Guanidine / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D011489:Protein Denaturation; D020673:Protein Renaturation; D015843:Serpins / Q000737:chemistry / Q000378:metabolism; D013696:Temperature; D000979:alpha-2-Antiplasmin / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism
chemical_list: D015843:Serpins; D000979:alpha-2-Antiplasmin; D019791:Guanidine