Apfel C M, Evers S, Hubschwerlen C, Pirson W, Page M G, Keck W
Pharma Research Basel, F. Hoffmann-La Roche Ltd., CH-4070 Basel, Switzerland.
Antimicrob Agents Chemother. 2001 Apr;45(4):1053-7. doi: 10.1128/AAC.45.4.1053-1057.2001.
An assay was developed to determine the activity of peptide deformylase (PDF) inhibitors under conditions as close as possible to the physiological situation. The assay principle is the detection of N-terminal [35S]methionine labeling of a protein that contains no internal methionine. If PDF is active, the deformylation of the methionine renders the peptide a substrate for methionine aminopeptidase, resulting in the removal of the N-terminal methionine label. In the presence of a PDF inhibitor, the deformylation is blocked so that the N-formylated peptide is not processed and the label is detected. Using this assay, it is possible to determine the PDF activity under near-physiological conditions in a cell-free transcription-translation system as well as in intact bacterial cells.
已开发出一种检测方法，用于在尽可能接近生理状况的条件下测定肽脱甲酰基酶（PDF）抑制剂的活性。该检测原理是检测不含内部甲硫氨酸的蛋白质的N端[35S]甲硫氨酸标记。如果PDF有活性，甲硫氨酸的脱甲酰基作用会使该肽成为甲硫氨酸氨肽酶的底物，导致N端甲硫氨酸标记被去除。在存在PDF抑制剂的情况下，脱甲酰基作用被阻断，因此N-甲酰化肽不会被加工，从而检测到该标记。使用这种检测方法，可以在无细胞转录-翻译系统以及完整细菌细胞中，在接近生理条件下测定PDF活性。
