Hong Hui, Samborskyy Markiyan, Usachova Katsiaryna, Schnatz Katharina, Leadlay Peter F
Department of Biochemistry, University of Cambridge, Cambridge CB2 1GA, UK.
Beilstein J Org Chem. 2017 Nov 13;13:2408-2415. doi: 10.3762/bjoc.13.238. eCollection 2017.
Clethramycin from DSM4137, and mediomycins (produced together with clethramycin from ), are near-identical giant linear polyenes apparently constructed from, respectively, a 4-guanidinobutanoate or 4-aminobutanoate starter unit and 27 polyketide extender units, and bearing a specific -sulfonate modification at the C-29 hydroxy group. We show here that mediomycins are actually biosynthesised not by use of a different starter unit but by direct late-stage deamidination of (desulfo)clethramycin. A gene () encoding a candidate sulfotransferase has been located in both gene clusters. Deletion of this gene in DSM4137 led to accumulation of desulfoclethramycin only, instead of a mixture of desulfoclethramycin and clethramycin. The mediomycin gene cluster does not encode an amidinohydrolase, but when three candidate amidinohydrolase genes from elsewhere in the genome were individually expressed in and assayed, only one of them (), located 670 kbp away from the mediomycin gene cluster on the chromosome, catalysed the removal of the amidino group from desulfoclethramycin. Subsequent cloning of into DSM4137 caused mediomycins A and B to accumulate at the expense of clethramycin and desulfoclethramycin, respectively, a rare case where an essential biosynthetic gene is not co-located with other pathway genes. Clearly, both desulfoclethramycin and clethramycin are substrates for this amidinohydrolase. Also, purified recombinant sulfotransferase from DSM4137, in the presence of 3'-phosphoadenosine-5'-phosphosulfate as donor, efficiently converted mediomycin B to mediomycin A in vitro. Thus, in the final steps of mediomycin A biosynthesis deamidination and sulfotransfer can take place in either order.
来自DSM4137的克来霉素以及麦迪霉素（与克来霉素一起产生）是几乎相同的巨型线性多烯，显然分别由4-胍基丁酸酯或4-氨基丁酸酯起始单元和27个聚酮链延伸单元构建而成，并在C-29羟基处带有特定的磺酸酯修饰。我们在此表明，麦迪霉素实际上并非通过使用不同的起始单元进行生物合成，而是通过（去硫）克来霉素的直接后期脱酰胺作用。在两个基因簇中都定位到了一个编码候选磺基转移酶的基因（）。在DSM4137中缺失该基因仅导致去硫克来霉素的积累，而不是去硫克来霉素和克来霉素的混合物。麦迪霉素基因簇不编码酰胺水解酶，但是当来自基因组其他位置的三个候选酰胺水解酶基因在中单独表达并进行检测时，其中只有一个（），位于染色体上距麦迪霉素基因簇670 kbp处，催化从去硫克来霉素上去除脒基。随后将克隆到DSM4137中导致麦迪霉素A和B分别积累，而克来霉素和去硫克来霉素减少，这是一个罕见的情况，即一个必需的生物合成基因与其他途径基因不在同一位置。显然，去硫克来霉素和克来霉素都是这种酰胺水解酶的底物。此外，来自DSM4137的纯化重组磺基转移酶在3'-磷酸腺苷-5'-磷酸硫酸作为供体的存在下，在体外有效地将麦迪霉素B转化为麦迪霉素A。因此，在麦迪霉素A生物合成的最后步骤中，脱酰胺作用和磺基转移可以以任意顺序发生。
