The contact zones in human alpha2-macroglobulin--functional domains important for the regulation of the trapping mechanism.
A functional domain termed the contact zone, which is the region of a subunit interacting with another non-covalently bound subunit, is suggested to play a decisive role in the trapping mechanism of human alpha2-macroglobulin. Tetrameric alpha2-macroglobulin can be dissociated into stable dimers with intact thiol esters by sodium thiocyanate, whereby the contact zones are disrupted. The dissociation leads to significant conformational changes, as studied by ultraviolet-difference spectroscopy, CD, fluorescence and affinity partitioning. The conformation of the dimers is similar to that of MeNH2-treated alpha2-macroglobulin, in which the thiol esters are cleaved, a conformational state with a closed trap occurs, and receptor-recognition sites are exposed. The receptor-binding domain is at least partly exposed in the dimer, as judged by binding of specific mAbs. The bait region in the dimers can be cleaved by proteases, and activation of the thiol esters ensues without binding of the protease. When the dimers were treated with MeNH2, no conformational changes could be detected by ultraviolet-difference spectroscopy or CD. The conformational changes occurring on dissociation into dimers are suggested to be related to trap closure and receptor-recognition-site exposure without cleavage of the thiol esters. The model presented here suggests that two separate conformational changes occur in alpha2-macroglobulin upon activation. The first involves changes at the contact zones as a result of the thiol-ester cleavage, and the second causes exposure of the receptor-recognition sites and closure of the trap.
Shanbhag
V P
VP
Department of Biochemistry, Umeå University, Sweden.
Stigbrand
T
T
Jensen
P E
PE
D000911:Antibodies, Monoclonal; D001665:Binding Sites; D002942:Circular Dichroism; D019281:Dimerization; D010450:Endopeptidases / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D015394:Molecular Structure; D011487:Protein Conformation; D011489:Protein Denaturation; D013050:Spectrometry, Fluorescence; D013056:Spectrophotometry, Ultraviolet; D013438:Sulfhydryl Compounds / Q000737:chemistry; D013861:Thiocyanates; D000511:alpha-Macroglobulins / Q000737:chemistry* / Q000276:immunology / Q000378:metabolism
D000911:Antibodies, Monoclonal; D013438:Sulfhydryl Compounds; D013861:Thiocyanates; D000511:alpha-Macroglobulins; C024553:sodium thiocyanate; D010450:Endopeptidases