Molécules de Communication et Adaptation des Microorganismes, UMR 7245 CNRS, Muséum National d'Histoire Naturelle, CP 54, 57 rue Cuvier, 75005 Paris, France.
Chembiochem. 2012 May 7;13(7):1046-52. doi: 10.1002/cbic.201200016. Epub 2012 Apr 5.
Microcin J25 is the archetype of a growing class of bacterial ribosomal peptides possessing a knotted topology (lasso peptides). It consists of an eight-residue macrolactam ring through which the C-terminal tail is threaded. It is biosynthesized as a precursor that is processed by two maturation enzymes (McjB/McjC). Insights into the mechanism of microcin J25 biosynthesis have been provided previously by mutagenesis of the precursor peptide in vivo. In this study we have demonstrated distinct functions of McjB and McjC in vitro for the first time, based on the detection of reaction intermediates. McjB was characterized as a new ATP-dependent cysteine protease, whereas McjC was confirmed to be a lactam synthetase. The two enzymes were functionally interdependent, likely forming a structural complex. Their substrate preference was directly investigated with the aid of mutated precursor peptides. Depending on the substitutions, microcin J25 variants with either a lasso or branched-cyclic topology could be generated in vitro.
微菌素 J25 是一类具有成环拓扑结构（套索肽）的新型核糖体肽的原型，这类核糖体肽还在不断增加。它由一个 8 个残基大环内酯环组成，C 末端尾部穿过该环。它作为前体被生物合成，然后由两种成熟酶（McjB/McjC）加工。先前通过体内突变前体肽，对微菌素 J25 生物合成机制有了初步了解。本研究首次基于反应中间产物的检测，体外证明了 McjB 和 McjC 的不同功能。McjB 被鉴定为一种新的 ATP 依赖性半胱氨酸蛋白酶，而 McjC 被证实为一种内酯合成酶。这两种酶在功能上相互依赖，可能形成一个结构复合物。借助突变前体肽直接研究了它们的底物偏好性。根据取代情况，微菌素 J25 的变体可以在体外生成套索或支链环状拓扑结构。
