Functional analysis of enzymes involved in the co-translational modification of nascent chains

新生多肽的N-末端加工是所有生物体中存在的重要蛋白水解途径。在细菌中，新生链在两个连续的反应中被处理。首先，肽脱甲酰基酶（PDF）去除N-末端甲酰基。其次，N-末端甲硫氨酸通过甲硫氨酸氨基肽酶（MAP）蛋白水解除去，该反应取决于倒数第二个氨基酸的性质。真核生物缺乏N-末端的乙酰化，新生链直接由MAP加工，但大多数新生链在其N-末端被核糖体相关的N-乙酰转移酶（NAT）乙酰化。酶促修饰发生在翻译的早期阶段，伴随着蛋白质折叠或生长的多肽共翻译靶向至易位子的初始步骤。为了保持这个过程的准确性，它必须高度协调。我们的科学目标是分析（i）PDF和MAP与新生链相互作用的进一步因素的时空协调（ii）细菌N-乙酰基转移酶和SspB中的衔接蛋白的潜在核糖体缔合，SspB是一种参与AAA+蛋白酶ClpXP识别和降解SsrA标记蛋白的蛋白质，以及（iii）N-末端加工和修饰对于新合成蛋白质的折叠和稳定性的重要性。

N-terminal processing of nascent polypeptides is an essential proteolytic pathway existing in all organisms. In bacteria, nascent chains are processed in two consecutive reactions. First, peptide deformylase (PDF) removes the N-terminal formyl group. Second, the N-terminal methionine is proteolytically removed by methionine aminopetidase (MAP), a reaction that depends on the nature of the penultimate amino acid. Eukaryotes lack N-terminal formylation and nascent chains are directly processed by MAPs, but most nascent chains are acetylated at their N-terminus by ribosome associated N-acetyltransferases (NATs). Enzymatic modifications occur in early phases of translation concomitantly with initial steps of protein folding or co-translational targeting of the growing polypeptide to the translocon. In order to maintain accuracy of this process, it must be highly coordinated. Our scientific aim is to analyze (i) the spatial and temporal coordination of PDF and MAP with further factors interacting with nascent chains (e.g. the chaperone trigger factor and the targeting factors SRP and SecA), (ii) the potential ribosome association of bacterial N-acetyltransferases and the adaptor prote in SspB, a protein involved in the recognition and degradation of SsrA-tagged proteins by the AAA+ protease ClpXP, and (iii) the importance of N-terminal processing and modification for folding and stability of newly synthesized proteins.