鱼肝中胆汁酸结合酶的纯化与特性分析

Purification and characterization of the enzymes of bile acid conjugation from fish liver.

Vessey D A, Benfatto A M, Zerweck E, Vestweber C

Liver Study Unit, Veterans Administration Medical Center, San Francisco, CA 94121.

Comp Biochem Physiol B. 1990;95(4):647-52. doi: 10.1016/0305-0491(90)90299-9.

Abstract

The two steps in bile acid conjugation have been studied in subcellular fractions of liver from three species of fish; vermillion rockfish, canary rockfish and ling codfish. 2. The bile acid: coenzyme A (CoA) ligase activity in homogenates and isolated microsomes is undetectable due to indeterminate factors. 3. A purification scheme is presented which eliminates the interfering factors. The purified ligase was found to have a lower affinity for bile acids as compared to the mammalian form and to be present in much lower titer. 4. Since it appears to be the rate controlling enzyme in all species, it is expected that the rate of bile acid conjugation is much slower in non-mammalian liver as compared to mammalian liver. 5. The bile acid-CoA:taurine N-acyltransferase was found to exist as a dimer of molecular weight 100,000, in contrast to the monomeric mammalian forms. 6. The only major kinetic difference is that the fish liver forms have rates of glycine conjugation which are only 1-2% of the rate with taurine, in part due to a very high Km for glycine.

摘要

对三种鱼类（朱红岩鱼、加那利岩鱼和太平洋鳕）肝脏的亚细胞组分中胆汁酸结合的两个步骤进行了研究。2. 由于不确定因素，匀浆和分离的微粒体中胆汁酸：辅酶A（CoA）连接酶活性无法检测到。3. 提出了一种纯化方案，该方案消除了干扰因素。发现纯化后的连接酶与哺乳动物形式相比，对胆汁酸的亲和力较低，且效价低得多。4. 由于它似乎是所有物种中控制速率的酶，预计与哺乳动物肝脏相比，非哺乳动物肝脏中胆汁酸结合的速率要慢得多。5. 发现胆汁酸-CoA：牛磺酸N-酰基转移酶以分子量为100,000的二聚体形式存在，这与单体形式的哺乳动物酶不同。6. 唯一主要的动力学差异是鱼肝形式的甘氨酸结合速率仅为牛磺酸结合速率的1-2%，部分原因是对甘氨酸的Km值非常高。