环缩肽cotransin被描述为通过Sec61蛋白传导通道的信号序列识别机制抑制一小部分蛋白的生物合成。然而,尚不清楚共转录素的选择性,即有多少蛋白质是敏感的。此外,尚未描述介导coransin敏感性的信号序列中的共有基序。为了解决这些问题,我们使用coransin处理的人肝癌细胞进行了蛋白质组学研究,并在细胞培养技术中通过氨基酸结合定量质谱进行了稳定同位素标记。我们使用饱和浓度的coransin(30微摩尔)来鉴定敏感性较低的蛋白质,并将后者与完全抗性的蛋白质区分开。我们发现,在这些条件下,几乎所有分泌蛋白的生物合成都对coransin敏感。相比之下,大多数完整膜蛋白的生物合成是抗coransin的。信号序列的共肽敏感性既与它们的长度无关,也与它们的疏水性无关。相反,在信号锚定序列的情况下,我们首次确定了介导coransin敏感性的构象共识基序。
The cyclodepsipeptide cotransin was described to inhibit the biosynthesis of a small subset of proteins by a signal sequence-discriminatory mechanism at the Sec61 protein-conducting channel. However, it was not clear how selective cotransin is, i.e. how many proteins are sensitive. Moreover, a
consensus motif in signal sequences mediating cotransin sensitivity has yet not been described. To address these questions, we performed a proteomic study using cotransin-treated human hepatocellular carcinoma cells and the stable isotope labelling by amino acids in cell culture technique in combination with quantitative mass spectrometry. We used a saturating concentration of cotransin (30 micromolar) to identify also less-sensitive proteins and to discriminate the latter from completely resistant proteins. We found that the biosynthesis of almost all secreted proteins was cotransin-sensitive under these conditions. In contrast, biosynthesis of the majority of the integral membrane proteins was cotransin-resistant. Cotransin sensitivity of signal sequences was neither related to their length nor to their hydrophobicity. Instead, in the case of signal anchor sequences, we identified for the first time a conformational
consensus motif mediating cotransin sensitivity.