Structural and functional characterization of Escherichia coli peptidyl-prolyl cis-trans isomerases.
Structural and functional characterization of Escherichia coli peptidyl-prolyl cis-trans isomerases.
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大肠杆菌肽基脯氨酰顺反异构酶的结构和功能表征。
DOI：
10.1111/j.1432-1033.1992.tb17002.x
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发表时间：
1992
期刊：
影响因子：
--
通讯作者：
Bächinger,HP
中科院分区：
文献类型：
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作者：
Compton,LA;Davis,JM;Macdonald,JR;Bächinger,HP
从大肠杆菌中纯化了肽基脯氨酰顺式-反式异构酶（PPIases），这种酶催化脯氨酸提供氮的肽键的顺式-反式异构化。在这种生物体中，至少存在两种PPIase。阳离子（周质）和阴离子（胞质）PPI酶均被aKi为25-50 μM的环孢菌素A抑制，该浓度比真核PPI酶所需的浓度高1000倍。虽然等电聚焦表明两种酶的等电点相差至少4.0 pH单位，但酶对四肽底物琥珀酰-Ala-Ala-Pro-Phe-甲基-香豆酰胺的比活性是相等的。两种酶对一系列取代的琥珀酰-Ala-Xaa-Pro-Phe帕拉对硝基苯胺四肽的活性表明，原核蛋白的活性位点的结构和功能与真核亲环蛋白的活性位点相似。两种酶都能够催化热变性III型胶原蛋白的重折叠。针对周质PPI酶的抗体不识别胞质酶，表明两种形式之间的表位存在显著差异。圆二色谱分析表明，该阳离子蛋白的二级结构由17%的α-螺旋、34%的β-折叠、17%的转角和33%的无规卷曲组成，与人胞浆PPIase非常相似。
Peptidyl‐prolylcis‐transisomerases (PPIases), enzymes that catalyze thecis‐transisomerization of peptide bonds to which proline contributes the nitrogen, were purified fromEscherichia coli. In this organism, at least two PPIases are present. Both the cationic (periplasmic) and anionic (cytoplasmic) PPIases are inhibited by cyclosporin A with aKiof 25–50 μM, a concentration 1000‐fold higher than that required for eukaryotic PPIases. Although isoelectric focusing indicates that the two enzymes differ in isoelectric point by at least 4.0 pH units, the specific activities of the enzymes toward the tetrapeptide substrate succinyl‐Ala‐Ala‐Pro‐Phe‐methyl‐coumarylamide are equivalent. The activity of both enzymes for a series of substituted succinyl‐Ala‐Xaa‐Pro‐Phe‐para‐nitroanilide tetrapeptides suggests that the structure and function of the active site of the prokaryotic proteins is similar to that of eukaryotic cyclophilins. Both enzymes are capable of catalyzing the refolding of thermally denatured type III collagen. Antibodies against the periplasmic PPIase do not recognize the cytoplasmic enzyme, indicating significant differences in epitopes between the two forms. Circular dichroism spectroscopy indicates that the secondary structure of the cationic protein consists of 17%α‐helix, 34%β‐sheet, 17% turns, 33% random coil and is very similar to human cytosolic PPIase.