Scholz C, Schindler T, Dolinski K, Heitman J, Schmid F X
Biochemisches Laboratorium, Universität Bayreuth, Germany.
FEBS Lett. 1997 Sep 1;414(1):69-73. doi: 10.1016/s0014-5793(97)00979-4.
The prolyl isomerase activity of cyclophilins is traditionally measured by an assay in which prolyl cis/trans isomerization in a chromogenic tetrapeptide is coupled with its isomer-specific cleavage by chymotrypsin. Two variants of mitochondrial cyclophilin with substitutions in the presumed active site (R73A and H144Q) are inactive in the protease-coupled assay, but show almost wild-type activity in an assay that is based on the catalysis of a proline-limited protein folding reaction. This prolyl isomerase assay is preferable, both because coupling with proteolysis is avoided and because an intact protein instead of a short peptide is used as a substrate. Possibly, some earlier conclusions about the catalytic mechanism and the involvement of the prolyl isomerase activity in the cellular function of immunophilins may need reevaluation.
亲环蛋白的脯氨酰异构酶活性传统上是通过一种测定方法来测量的，在该方法中，生色四肽中的脯氨酰顺/反异构化与其被胰凝乳蛋白酶的异构体特异性切割相偶联。线粒体亲环蛋白的两个变体在假定的活性位点有取代（R73A和H144Q），在蛋白酶偶联测定中无活性，但在基于脯氨酸限制的蛋白质折叠反应催化的测定中显示出几乎野生型的活性。这种脯氨酰异构酶测定更可取，这既是因为避免了与蛋白水解的偶联，也是因为使用完整的蛋白质而不是短肽作为底物。可能，一些关于催化机制以及脯氨酰异构酶活性在亲免素细胞功能中的参与的早期结论可能需要重新评估。
