Symmetry of the inhibitory unit of human alpha 2-macroglobulin.
Human alpha 2-macroglobulin (alpha 2M) of Mr approximately 720,000 is a proteinase inhibitor whose four identical subunits are arranged to form two adjacent inhibitory units. At present, the spatial arrangement of the two subunits which form one inhibitory unit (the functional "half-molecule") is not known. Treatment of alpha 2M with either 0.5 mM dithiothreitol (DTT) or 4 M urea results in dissociation of the native tetramer into two half-molecules of Mr approximately 360,000. These half-molecules retain trypsin inhibitory activity, but in each case, the reaction results in reassociation of the half-molecules to produce tetramers of Mr approximately 720,000. However, when reacted with plasmin, the preparations of half-molecules have different properties. DTT-induced half-molecules protect the activity of plasmin from inhibition by soybean trypsin inhibitor (STI) without reassociation, while urea-induced half-molecules show no ability to protect plasmin from reaction with STI. High-performance size-exclusion chromatography and sedimentation velocity ultracentrifugation studies were then used to estimate the Stokes radius (Re) of alpha 2M and both DTT- and urea-induced half-molecules of alpha 2M. The Re of tetrameric alpha 2M was 88-94 A, while that of DTT-induced half-molecules was 57-60 A and urea-induced half-molecules 75-77 A. These results demonstrate that DTT- and urea-induced half-molecules have fundamentally different molecular dimensions as well as inhibitory properties. The hydrodynamic data suggest that the urea-induced half-molecule is a "rod"-like structure, although it is not possible to predict the three-dimensional structure of this molecule with the available data.(ABSTRACT TRUNCATED AT 250 WORDS)
Roche
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PA
Department of Pathology, Duke University Medical Center, Durham, North Carolina 27710.
Salvesen
G S
GS
Pizzo
S V
SV
D004229:Dithiothreitol; D005341:Fibrinolysin / Q000037:antagonists & inhibitors; D006801:Humans; D066298:In Vitro Techniques; D008970:Molecular Weight; D011487:Protein Conformation; D014361:Trypsin Inhibitors; D014508:Urea; D000511:alpha-Macroglobulins / Q000494:pharmacology
D014361:Trypsin Inhibitors; D000511:alpha-Macroglobulins; D014508:Urea; D005341:Fibrinolysin; D004229:Dithiothreitol