摘要：

Circular dichroism spectroscopy, sedimentation velocity and ultraviolet difference spectroscopy were used to compare α 2-macroglobulin, α 2-macroglobulin-trypsin complex and α 2-macroglobulin-methylamine complex. The circular dichroic spectrum of native α 2-macroglobulin is significantly changed in shape and magnitude following reaction with either trypsin or methylamine. The spectra of α 2-macroglobulin-trypsin and α 2-macroglobulin-methylamine are, however, indistinguishable. The ultraviolet difference spectrum between α 2-macroglobulin-methylamine and native α 2-macroglobulin displays a tyrosine blue shift consistent with the exposure of several tyrosine residues to solvent. The conformational change which occurs in α 2-macroglobulin during reaction with methylamine follows pseudo-first-order kinetics. T 1/2 was 10.5 min for the reaction with 200 mM methylamine at pH 8.0 and 45 min for the reaction with 50 mM methylamine, also at pH 8.0. Reaction of methylamine with α 2-macroglobulin results in loss of trypsin-binding activity which appears to be a direct consequence of the conformational change induced by methylamine. A sedimentation coefficient (s o 20,w) of 20.5 was determined for α 2-macroglobulin-methylamine compared to a value of 18.5 for unreacted α 2-macrogiobulin. This increase in sedimentation velocity is attributed to a 10% decrease in α 2-macroglobulin Stokes radius, α 2-Macroglobulin-trypsin complex prepared by reaction of the protease ata 2-fold molar excess with the inhibitor has a s o 20,w of 203. Although this sedimentation coefficient does reflect compacting of the α 2-macroglobulin structure compared to native α 2-macroglobulin, it is not large enough to rule out significant protrusion of the proteases from within pockets in the α 2-macroglobulin structure.

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