Purification and properties of tryptophan 5-monooxygenase from rat brain-stem.
Purification and properties of tryptophan 5-monooxygenase from rat brain-stem.
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大鼠脑干色氨酸5-单加氧酶的纯化及其性质。
DOI：
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发表时间：
1982
期刊：
影响因子：
--
通讯作者：
H. Fujisawa
中科院分区：
文献类型：
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作者：
H. Nakata;H. Fujisawa
使用 Sepharose CL-6B、DEAE-Sepharose CL-6B 和蝶啶-琼脂糖色谱法，从大鼠脑干中纯化色氨酸 5-单加氧酶约 5,500 倍，达到表观同质性，30 ℃ 下比活性为 374 nmol min-1 mg-1。两种不同的活性形式可通过 DEAE-Sepharose CL-6B 分离，并根据它们从凝胶柱中的洗脱顺序指定为形式 I 和形式 II。通过Ultrogel AcA 34上的凝胶过滤测定形式I的表观分子量为300,000，通过梯度聚丙烯酰胺凝胶电泳测定为288,000。该酶在十二烷基硫酸钠/聚丙烯酰胺凝胶电泳上显示一条单条带，其分子量估计为59,000，表明该酶可能由四个相同的亚基组成。使用辛二酰亚胺酸二甲酯作为双功能试剂进行交联研究，进一步表明了酶的四聚体结构。添加 Fe2+ 可使酶活性提高约 3.5 倍。动力学研究表明，这种激活与 V 值的增加有关。纯化的酶具有苯丙氨酸羟基化活性，但不具有酪氨酸羟基化活性。
Tryptophan 5-monooxygenase was purified approximately 5,500-fold, to apparent homogeneity with a specific activity of 374 nmol min-1 mg-1 at 30 degrees C, from rat brain-stem using Sepharose CL-6B, DEAE-Sepharose CL-6B and pteridine-agarose chromatography. Two distinct active forms were separable by DEAE-Sepharose CL-6B and designated as form I and form II based on their order of elution from the gel column. The apparent molecular weight of form I was determined to be 300,000 by gel filtration on Ultrogel AcA 34 and 288,000 by gradient polyacrylamide gel electrophoresis. The enzyme gave a single band on sodium dodecylsulfate/polyacrylamide gel electrophoresis, the molecular weight of which was estimated to be 59,000, indicating that the enzyme might be composed of four identical subunits. The tetrameric structure of the enzyme was further suggested by cross-linking studies using dimethyl suberimidate as a bifunctional reagent. The enzyme activity was stimulated approximately 3.5-fold by the addition of Fe2+. Kinetic studies revealed that this activation was associated with an increase of V value. The purified enzyme had an activity of phenylalanine hydroxylation but not an activity of tyrosine hydroxylation.
通过亲和层析纯化大鼠肝苯丙氨酸羟化酶。
影响因子：
3.9
作者：
Al-Janabi,JM
通讯作者：
Al-Janabi,JM