Purification and Characteriaztion of the alpha 2 Macroglobulin

Purification and Characteriaztion of the alpha 2 Macroglobulin Protease Inhibitor from the Rana trigrin n The alpha 2 Macroglobulin (A 2 M) are classified as broadspectrum inhibitorsbecause of their ability to entrap protease of different specificites and cata-lytic class. In the previous studies from our laboratory, the putative frog A 2 Mc. DNA containing thioester region sequences has been cloned. In this report, the. A 2 M-like protease inhibitor from plasma of the Rana trigrina was purified to a-pparent homogeneity by DEAE-sepharose CL-6 B anion-exchange column and preparat-ive electrophoresis. The purified protein resembled verterbrate A 2 Ms in that itdisplays a broad specificity and inhibited the activity of trypsin, chymotryps-in, papain, pepsin and elastase. Sensitivity of this purified protein to methy-lamine and autolytic cleavage suggested the presence of an internal bata-cyste-inyl-r-glutamyl thioester. The purified protein was tetramer of 180 KDa and had and apparent mass of approximately 720 Kda when examined by nonreducing and re-ducing sodium dodecylsulfate-polyacrylamide gel (SDS-PAGE), and gel filteration. When boiled in SDS it under went autolytic fragmentation to produce two bandsof approximately 110 and 80 KDa resemble to human A 2 M. The amino acid compositi-on showed similarities with the wellcharacterized vertebrate A 2 Ms. These resu-lts suggested the purified protein was an A 2 M-like protease inhibitor.