Characterisation and exploitation of a promiscuous non-ribosomal peptide cyclase

大多数临床使用的抗生素来自链霉菌属和其他密切相关的土壤细菌产生的天然产物。这些药物最初是在1940-1960年抗生素发现的“黄金时代”被发现并引入临床的。在过去的半个世纪里，由于滥用，这些药剂的效用已经受到侵蚀。因此，现在迫切需要发现新的抗生素来治疗耐药性细菌感染。对抗菌剂耐药性的日益关注，加上未能从筛选合成化合物的大型库中找到新的线索，导致人们对天然产物发现重新产生兴趣。不幸的是，绝大多数的微生物还没有被培养出来，而对于那些已经被培养出来的微生物来说，只有一小部分的天然产物是在实验室中产生的。克服这一问题的常规方法通常依赖于对生产生物体进行耗时的遗传修饰或使用知之甚少的“激发子”化合物来启动生产。这种方法的瓶颈是，大量的时间可能花费在一个生物合成途径上，其产物可能永远不会产生或不是抗生素。此外，即使成功地激活了抗生素途径，发现先导化合物也只是药物发现的第一阶段。许多抗生素是从一类微生物的天然产物称为非核糖体肽。一旦确定了一个令人兴奋的前景，未来的发展最终取决于其可访问性。微生物发酵很少提供足够的化合物来向前推进，因此化学合成通常用于生产测试发现线索的临床潜力所需的数量，重要的是，化学多样性。这里的问题是，绝大多数非核糖体肽是环状的，并且环化反应通常非常成问题，并且产生低收率的最终化合物。在自然界中，环化反应是通过生物合成途径的一部分进行的，称为硫酯酶结构域。我们最近发现了一种新的环化酶，这是混杂的肽底物，它环化。这是令人兴奋的，我们想了解这种酶是如何工作的，这样我们就可以利用它的生物技术潜力。例如，改进抗生素的化学合成。我们相信，这最终可以帮助更多的药物进入诊所，可能使它们更便宜，更广泛地获得。

The majority of clinically used antibiotics are derived from natural products produced by Streptomyces species and other closely related soil bacteria. These drugs were primarily discovered and introduced into the clinic during a 'golden era' of antibiotic discovery that spanned 1940-1960. The utility of these agents has been eroded over the last half-century due to misuse. As a consequence, there is now an urgent need to discover new antibiotics to treat drug resistant bacterial infections. Growing concerns about resistance to antibacterial agents combined with the failure to find new leads from the screening of large libraries of synthetic compounds has led to a renewed interest in natural products discovery. Unfortunately, the overwhelming majority of microbes have yet to be cultured, and for those that have, only a small fraction of their natural products are produced in the laboratory. Conventional approaches to overcome this problem, typically rely upon time consuming genetic modification of the producing organism or the use of poorly understood 'elicitor' compounds to switch on production. The bottleneck with this approach is the fact that a large amount time can be spent on one biosynthetic pathway whose product could never be produced or is not an antibiotic. Moreover, even if success in activating an antibiotic pathway is achieved, discovery of a lead compound is only the first stage of drug discovery. Many antibiotics are derived from a class of microbial natural products called non-ribosomal peptides. Once an exciting prospect is identified, future development is ultimately dictated by its accessibility. Microbial fermentation rarely provides sufficient compound to move forward and therefore chemical synthesis is typically used to produce the quantity, and importantly, the chemical diversity of analogues necessary for testing the clinical potential of a discovery lead. The problem here is that the vast majority of non-ribosomal peptides are cyclic and cyclisation reactions are typically very problematic and produce a low yield of the final compound. In nature, the cyclisation reaction is carried out by a part of the biosynthetic pathway called a thioesterase domain. We recently identified a novel cyclase enzyme, which is promiscuous with respect to the peptide substrates it cyclises. This is exciting and we want to understand how this enzyme works so we can harness its potential for biotechnology. For example, to improve chemical synthesis of antibiotics. We believe this could ultimately help more medicines reach the clinic, possibly making them less expensive and more widely available.