Physical properties of human α2-macroglobulin following reaction with methylamine and trypsin

doi:10.1016/0167-4838(82)90252-7

Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology

Volume 705, Issue 3, 10 August 1982, Pages 306-314

Biochimica et Biophy...

https://doi.org/10.1016/0167-4838(82)90252-7 Get rights and content

Abstract

Circular dichroism spectroscopy, sedimentation velocity and ultraviolet difference spectroscopy were used to compare α₂-macroglobulin, α₂-macroglobulin-trypsin complex and α₂-macroglobulin-methylamine complex. The circular dichroic spectrum of native α₂-macroglobulin is significantly changed in shape and magnitude following reaction with either trypsin or methylamine. The spectra of α₂-macroglobulin-trypsin and α₂-macroglobulin-methylamine are, however, indistinguishable. The ultraviolet difference spectrum between α₂-macroglobulin-methylamine and native α₂-macroglobulin displays a tyrosine blue shift consistent with the exposure of several tyrosine residues to solvent. The conformational change which occurs in α₂-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 α₂-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^o₂₀,w) of 20.5 was determined for α₂-macroglobulin-methylamine compared to a value of 18.5 for unreacted α₂-macrogiobulin. This increase in sedimentation velocity is attributed to a 10% decrease in α₂-macroglobulin Stokes radius, α₂-Macroglobulin-trypsin complex prepared by reaction of the protease ata 2-fold molar excess with the inhibitor has a s^o₂₀,w of 203. Although this sedimentation coefficient does reflect compacting of the α₂-macroglobulin structure compared to native α₂-macroglobulin, it is not large enough to rule out significant protrusion of the proteases from within pockets in the α₂-macroglobulin structure.

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Physical properties of human α2-macroglobulin following reaction with methylamine and trypsin

Abstract

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

Biochim. Biophys. Acta

Biochem. Biophys. Res. Commun.

Clin. Chim. Acta

FEBS Lett.

Clin. Chim. Acta

J. Biol. Chem.

J. Biol. Chem.

Biochem. Biophys. Res. Commun.

Biochem. J.

J. Clin. Invest.

Biochem. J.

Biochem. J.

Physical properties of human α₂-macroglobulin following reaction with methylamine and trypsin