Tryptophan Synthase from Escherichia coli
Tryptophan Synthase from Escherichia coli
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来自大肠杆菌的色氨酸合酶
DOI：
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发表时间：
2013
期刊：
影响因子：
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通讯作者：
R. Jaenicke
中科院分区：
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作者：
T. Seifert;P. Bartholmes;R. Jaenicke
该/？2-大肠杆菌色氨酸合酶二聚体与8-苯胺基萘磺酸（ANS）的结合较弱。滴定染料在0.2 mM浓度与apo-/？2-大气压下的二聚体导致在480 nm（λ exc = 380 nm）处的荧光发射增加，这对应于配体与疏水残基的非特异性结合。增加静水压力影响AN S结合。高达700巴，f ANS荧光的S形增加反映了疏水表面积的增加，这可能是由亚基解离引起的。在~ 1 kbar时，达到最大值;超过该值，压力与配体结合竞争，导致荧光发射再次减少。压力释放导致急剧的荧光增强，归因于ANS结合到部分和可逆变性的酶。绘制总荧光增强对压力的曲线，其与通过亚基杂交监测的压力依赖性二聚体→单体转变平行（T. Seifert，P. Bartholmes，and R. Jaenicke，Biochemistry，in press）.
The /?2-dimer o f tryptophan synthase from Escherichia coli exhibits weak binding o f 8-anilinonaphthalene-lsulfonic acid (ANS). Titrating the dye at 0.2 mM concen tration with the apo-/?2-dimer at atmospheric pressure causes increased fluorescence emission at 480 nm (/.exc = 380 nm), corresponding to unspecific binding o f the ligand to hydrophobic residues. Increasing hydrostatic pressure affects AN S binding. Up to 700 bar, a sigmoidal increase o f ANS fluorescence reflects an increase in hydrophobic surface area, probably caused by subunit dissociation. At ~ 1 kbar, a maximum is reached; beyond this value, pressure com petes with ligand binding causing fluorescence emission to be de creased again. Pressure release leads to a drastic fluorescence enhance ment, ascribed to ANS binding to the partially and reversibly denatured enzyme. Plotting the total fluorescence enhancement vs. pressure yields a profile which parallels the pressure dependent dimer ^ monomer transition monitored by subunit hybridization (T. Seifert, P. Bartholmes, and R. Jaenicke, Biochemistry, in press).