High-efficiency incorporation in vivo of tyrosine analogues with altered hydroxyl acidity in place of the catalytic tyrosine-14 of Δ5-3-ketosteroid isomerase of Comamonas (Pseudomonas) testosteroni:: Effects of the modifications on isomerase kinetics
High-efficiency incorporation in vivo of tyrosine analogues with altered hydroxyl acidity in place of the catalytic tyrosine-14 of Δ5-3-ketosteroid isomerase of Comamonas (Pseudomonas) testosteroni:: Effects of the modifications on isomerase kinetics
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DOI：
10.1021/bi980454x
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发表时间：
1998-07-07
期刊：
影响因子：
2.9
通讯作者：
Benisek, WF
中科院分区：
文献类型：
--
作者：
Brooks, B;Phillips, RS;Benisek, WF
在大肠杆菌宿主中表达了Y55 F/Y88 F修饰形式的δ（δ）-3-酮甾体异构酶，其中活性位点酪氨酸-14被2-氟酪氨酸、3-氟酪氨酸和2，3-二氟酪氨酸（它们的酚羟基具有逐渐增大的酸性的氨基酸）取代，并纯化至高度均一性。测定了Y55 F/Y88 F KSI及其氟酪氨酸修饰物的稳态动力学性质。的结果的机制的影响，并进行了讨论。
Versions of the Y55F/Y88F modified form of Delta(5)-3-ketosteroid isomerase in which the active-site tyrosine-14 is replaced by 2-fluorotyrosine, 3-fluorotyrosine, and 2,3-difluorotyrosine, amino acids having progressively greater acidity of their phenolic hydroxyls, have been expressed in an Escherichia coli host and purified to high homogeneity. The steady-state kinetic properties of Y55F/Y88F KSI and its fluorotyrosine modified forms have been determined. The mechanistic implications of the results are presented and discussed.