Balibar Carl J, Garneau-Tsodikova Sylvie, Walsh Christopher T
Department of Biological and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA.
Chem Biol. 2007 Jun;14(6):679-90. doi: 10.1016/j.chembiol.2007.05.007.
The last stages of assembly of the aminocoumarin antibiotics, clorobiocin and coumermycin A(1), which target the GyrB subunits of bacterial DNA gyrase, involve enzymatic transfer of the pyrrolyl-2-carbonyl acyl group from a carrier protein (CloN1/CouN1) to the 3'-OH of the noviosyl moiety of the antibiotic scaffold. The enzyme, CouN7, will catalyze both the forward and back reaction on both arms of the coumermycin scaffold. This occurs via an O-acyl-Ser(101)-CouN7 intermediate, as shown by transient labeling of the enzyme with [(14)C]acetyl-S-CouN1 as donor and by inactivating mutation of the active site, Ser(101), to Ala. The intermediacy of the pyrrolyl-2-carbonyl-O-CouN7 allows net pyrrole transfer between distinct aminocoumarin scaffolds, for example, between the descarbamoylnovobiocin scaffold and coumermycin A(1) and vice versa. CouN7 also allows shuttling of surrogate acyl groups between noviosyl-aminocoumarin scaffolds to generate new antibiotic variants.
氨基香豆素类抗生素氯新生霉素和香豆霉素A（1）的组装最后阶段靶向细菌DNA促旋酶的GyrB亚基，这涉及将吡咯基-2-羰基酰基从载体蛋白（CloN1/CouN1）酶促转移至抗生素支架新霉糖部分的3'-OH。CouN7酶将催化香豆霉素支架双臂上的正向和反向反应。这通过O-酰基-Ser（101）-CouN7中间体发生，用[(14)C]乙酰-S-CouN1作为供体对该酶进行瞬时标记以及将活性位点Ser（101）突变为Ala使其失活均证明了这一点。吡咯基-2-羰基-O-CouN7中间体允许在不同的氨基香豆素支架之间进行净吡咯转移，例如在去氨甲酰基新霉素支架和香豆霉素A（1）之间，反之亦然。CouN7还允许在新霉糖基-氨基香豆素支架之间穿梭替代酰基以生成新的抗生素变体。
