title: Alpha 2-antiplasmin Enschede is not an inhibitor, but a substrate, of plasmin.
abstract: alpha 2-Antiplasmin Enschede is a variant of alpha 2-antiplasmin which has lost its ability to inhibit plasmin irreversibly and which is associated with a haemorrhagic disorder [Kluft et al. (1987) J. Clin. Invest. 80, 1391-1400]. The abnormal protein was purified from the plasma of a homozygous patient and subjected to one-dimensional peptide mapping using papain for digestion. A slightly abnormally migrating polypeptide (Mr 17,000) was found which represented the C-terminal part of the molecule (the N-terminus of the polypeptide corresponded to Gly-338 in normal alpha 2-antiplasmin) and which contained the reactive centre. The interaction of plasmin with alpha 2-antiplasmin Enschede was studied by adding plasmin to plasma of the homozygous patient. SDS/polyacrylamide-gel electrophoresis and immunoblotting showed that no complex persisted, but that the abnormal alpha 2-antiplasmin was cleaved into two fragments of Mr 56,000 and 14,000 respectively. The latter fragment co-migrated with the post-complex peptide, which is cleaved from normal alpha 2-antiplasmin during complex-formation with plasmin. In a purified system, catalytic amounts of plasmin rapidly cleaved alpha 2-antiplasmin Enschede into the aforementioned fragments. In kinetic studies alpha 2-antiplasmin Enschede reversibly and temporarily inhibited the plasmin-catalysed hydrolysis of D-valyl-L-leucyl-L-lysine p-nitroanilide ('S-2251') as a competitive inhibitor (Ki,app. 35 nM). It was concluded that alpha 2-antiplasmin Enschede apparently forms a normal complex with plasmin. The complex is, however, not stable, but disintegrates rapidly to a cleaved form of alpha 2-antiplasmin Enschede and active plasmin. The abnormal protein thus behaves like a substrate, instead of an inhibitor, of plasmin.
authors.affiliation: Gaubius Institute TNO, Leiden, The Netherlands.
authors.forename: D C
authors.initials: DC
authors.lastname: Rijken
authors.forename: E
authors.initials: E
authors.lastname: Groeneveld
authors.forename: C
authors.initials: C
authors.lastname: Kluft
authors.forename: H K
authors.initials: HK
authors.lastname: Nieuwenhuis
mesh_terms: D000595:Amino Acid Sequence; D004591:Electrophoresis, Polyacrylamide Gel; D005341:Fibrinolysin / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006474:Hemorrhagic Disorders / Q000097:blood*; D006801:Humans; D006868:Hydrolysis; D015151:Immunoblotting; D007700:Kinetics; D008969:Molecular Sequence Data; D009842:Oligopeptides / Q000378:metabolism; D010449:Peptide Mapping; D000979:alpha-2-Antiplasmin / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D009842:Oligopeptides; C053817:alpha(2)-antiplasmin Enschede; D000979:alpha-2-Antiplasmin; C018788:valyl-leucyl-lysine 4-nitroanilide; D005341:Fibrinolysin