Purification and Characteriaztion of the alpha 2 Macroglobulin Protease Inhibitor from the Rana trigrin

• Main Authors: Wu, Wen-Shin, 吳玟欣
• Other Authors: Lai Jing-Erh
• Format: Others
• Language: zh-TW
• Published: 1997
• Online Access: http://ndltd.ncl.edu.tw/handle/70315415223508181866

碩士 === 台北醫學院 === 醫學研究所 === 85 === The alpha 2 Macroglobulin (A2M) are classified as broad- spectrum inhibitorsbecause of their ability to entrap protease of different specificites and cata-lytic class. In the previous studies from our laboratory, the putative frog A2McDNA containing thioester region sequences has been cloned. In this report, theA2M-like protease inhibitor from plasma of the Rana trigrina was purified to a-pparent homogeneity by DEAE-sepharose CL-6B anion-exchange column and preparat-ive electrophoresis. The purified protein resembled verterbrate A2Ms 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 180KDa and had and apparent mass of approximately 720Kda when examined by non- reducing 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 80KDa resemble to human A2M. The amino acid compositi-on showed similarities with the well- characterized vertebrate A2Ms. These resu-lts suggested the purified protein was an A2M-like protease inhibitor.