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用Triton X-100和胰蛋白酶处理并结合硫酸铵分段盐析和多种柱层析,从正常小鼠肾脏纯化了芳香酰胺酶(Arylamidase、简写AAD,E.C.3.4.11.2.)。纯化的AAD经聚丙烯酰胺凝胶圆盘电泳(PAGE)鉴定为一条带。比活性为156μ/mg蛋白质。Sephadex G-200凝胶过滤测得分子量为213,000。十二烷基硫酸钠-聚丙烯酰胺凝胶圆盘电泳(SDS-PAGE)测得纯化的AAD分子是由二个相同亚基组成的二聚体。含糖量为14.0％。以L-亮氨酰-β-萘胺为底物时的K_m值为9.5×10~(-5)M,最适pH为7.0。纯化AAD受Co++激活以及受几种氨基酸尤其是非极性大侧链氨基酸抑制。
Arylamidase (AAD, E.C. 3.4.11.2.) Was purified from normal mouse kidneys by treatment with Triton X-100 and trypsin in combination with fractional salting-out with ammonium sulfate and various column chromatographies. Purified AAD was identified as a band by polyacrylamide gel electrophoresis (PAGE). Specific activity was 156μ / mg protein. Sephadex G-200 gel filtration yielded a molecular weight of 213,000. The purified AAD molecule as measured by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) is a dimer consisting of two identical subunits. The sugar content was 14.0%. The K_m value of L-leucyl-β-naphthylamine as a substrate was 9.5 × 10 -5 M, and the optimum pH was 7.0. Purified AAD is activated by Co ++ and is inhibited by several amino acids, especially nonpolar side chains.