Characterisation and exploitation of a promiscuous non-ribosomal peptide cyclase

临床上使用的抗生素大部分来源于链霉菌和其他密切相关的土壤细菌产生的天然产物。这些药物主要是在抗生素发现的“黄金时代”（1940-1960年）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被削弱了。因此，现在迫切需要发现新的抗生素来治疗耐药细菌感染。对抗菌剂耐药性的担忧日益增加，再加上未能从大型合成化合物文库中找到新的线索，这使得人们对天然产物的发现重新产生了兴趣。不幸的是，绝大多数微生物还没有被培养出来，对于那些已经培养出来的微生物，只有一小部分自然产物是在实验室里生产出来的。克服这一问题的传统方法通常依赖于对生产生物体进行耗时的基因改造，或者使用人们知之甚少的“激发子”化合物来启动生产。这种方法的瓶颈在于，在一种生物合成途径上可能花费大量时间，而这种途径的产物可能永远不会被生产出来，或者不是抗生素。此外，即使成功激活了抗生素途径，先导化合物的发现也只是药物发现的第一阶段。许多抗生素来源于一类被称为非核糖体肽的微生物天然产物。一旦确定了令人兴奋的前景，未来的发展最终取决于其可达性。微生物发酵很少提供足够的化合物，因此通常使用化学合成来生产数量，重要的是，化学多样性的类似物需要测试一个发现的临床潜力。这里的问题是，绝大多数非核糖体肽是环状的，而环化反应通常是非常有问题的，并且产生的最终化合物的产量很低。在自然界中，环化反应是由称为硫酯酶结构域的生物合成途径的一部分进行的。我们最近发现了一种新的环化酶，它对肽底物的环化是混杂的。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在生物技术上的潜力。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.