School of Biological Sciences, University of Auckland, Auckland, New Zealand.
AgResearch Limited, Food System Integrity, Palmerston North, New Zealand; Riddet Institute, Massey University, Palmerston North, New Zealand.
J Biol Chem. 2021 Jan-Jun;296:100432. doi: 10.1016/j.jbc.2021.100432. Epub 2021 Feb 18.
Nonribosomal peptide synthetases (NRPSs) are multimodular enzymes that produce a wide range of bioactive peptides, such as siderophores, toxins, and antibacterial and insecticidal agents. NRPSs are dynamic proteins characterized by extensive interdomain communications as a consequence of their assembly-line mode of synthesis. Hence, crystal structures of multidomain fragments of NRPSs have aided in elucidating crucial interdomain interactions that occur during different steps of the NRPS catalytic cycle. One crucial yet unexplored interaction is that between the reductase (R) domain and the peptide carrier protein (PCP) domain. R domains are members of the short-chain dehydrogenase/reductase family and function as termination domains that catalyze the reductive release of the final peptide product from the terminal PCP domain of the NRPS. Here, we report the crystal structure of an archaeal NRPS PCP-R didomain construct. This is the first NRPS R domain structure to be determined together with the upstream PCP domain and is also the first structure of an archaeal NRPS to be reported. The structure reveals that a novel helix-turn-helix motif, found in NRPS R domains but not in other short-chain dehydrogenase/reductase family members, plays a major role in the interface between the PCP and R domains. The information derived from the described PCP-R interface will aid in gaining further mechanistic insights into the peptide termination reaction catalyzed by the R domain and may have implications in engineering NRPSs to synthesize novel peptide products.
非核糖体肽合成酶（NRPSs）是一类多模块酶，能够产生广泛的生物活性肽，如铁载体、毒素、抗菌和杀虫剂。NRPSs 是动态蛋白，其特征是由于其装配线合成模式而导致的结构域间广泛的通讯。因此，NRPS 多结构域片段的晶体结构有助于阐明在 NRPS 催化循环的不同步骤中发生的关键结构域间相互作用。一个关键但尚未探索的相互作用是还原酶（R）结构域和肽载体蛋白（PCP）结构域之间的相互作用。R 结构域是短链脱氢酶/还原酶家族的成员，作为终止结构域发挥作用，可催化从 NRPS 的末端 PCP 结构域还原释放最终的肽产物。在这里，我们报告了一个古菌 NRPS PCP-R 双结构域构建体的晶体结构。这是第一个与上游 PCP 结构域一起确定的 NRPS R 结构域结构，也是第一个报道的古菌 NRPS 结构。该结构表明，在 NRPS R 结构域中发现的但在其他短链脱氢酶/还原酶家族成员中不存在的新型螺旋-转角-螺旋基序在 PCP 和 R 结构域之间的界面中起主要作用。所描述的 PCP-R 界面提供的信息将有助于深入了解 R 结构域催化的肽终止反应的机制，并可能对工程 NRPS 以合成新型肽产物具有重要意义。
