弗吉尼亚负鼠的腺苷脱氨酶同工酶：初步色谱分析和动力学研究。

Adenosine deaminase isoenzymes of the opossum Didelphis virginiana: initial chromatographic and kinetic studies.

作者信息

Niedzwicki J G, Liou C, Abernethy D R, Lima J E, Hoyt A, Lieberman M, Bethlenfalvay N C

机构信息

Oncology Center, North Dartmouth, MA 02747, USA.

出版信息

Comp Biochem Physiol B Biochem Mol Biol. 1995 Jun;111(2):291-8. doi: 10.1016/0305-0491(94)00249-t.

DOI:10.1016/0305-0491(94)00249-t

PMID:7599990

Abstract

Extracts of liver and spleen were used to isolate opossum adenosine deaminase isoenzymes (ADA1 and ADA2) and to determine their activities with adenosine and 2'-deoxyadenosine as substrates. Km values (microM) for adenosine and 2'-deoxyadenosine, respectively, as substrates for partially purified opossum liver adenosine deaminase isoenzymes were ADA1: 57 +/- 7 vs. 26 +/- 4 and ADA2: 285 +/- 25 vs. 580 +/- 92. In crude spleen extract, ADA2 activity was stable at 56 degrees C during 40 min of incubation. ADA1 activity declined in a linear fashion under the above conditions with an apparent T1/2 of 80 min. Sephadex G-150 column chromatography of crude spleen extract showed the apparent molecular weight of the ADA activity not inhibited by (+/-)-EHNA (i.e. ADA2) to be 170 kDa; ADA activity fully inhibited by (+/-)-EHNA (i.e. ADA1) eluted in the fractions corresponding to a molecular weight of 35 kDa.

摘要

使用肝脏和脾脏提取物来分离负鼠腺苷脱氨酶同工酶（ADA1和ADA2），并以腺苷和2'-脱氧腺苷作为底物测定它们的活性。作为部分纯化的负鼠肝脏腺苷脱氨酶同工酶底物的腺苷和2'-脱氧腺苷的Km值（微摩尔）分别为：ADA1：57±7对26±4，ADA2：285±25对580±92。在粗脾脏提取物中，ADA2活性在56℃孵育40分钟期间保持稳定。在上述条件下，ADA1活性呈线性下降，表观半衰期为80分钟。粗脾脏提取物的Sephadex G - 150柱色谱显示，不受（±）-EHNA抑制的ADA活性（即ADA2）的表观分子量为170 kDa；被（±）-EHNA完全抑制的ADA活性（即ADA1）在对应于分子量35 kDa的级分中洗脱。