Department of Chemistry and Chemical Biology, Harvard University, Cambridge, Massachusetts 02138, USA.
J Am Chem Soc. 2009 Dec 30;131(51):18230-1. doi: 10.1021/ja908916z.
Three periplasmic N-acetylmuramoyl-l-alanine amidases are critical for hydrolysis of septal peptidoglycan, which enables cell separation. The amidases cleave the amide bond between the lactyl group of muramic acid and the amino group of l-alanine to release a peptide moiety. Cell division amidases remain largely uncharacterized because substrates suitable for studying them have not been available. Here we have used synthetic peptidoglycan fragments of defined composition to characterize the catalytic activity and substrate specificity of the important Escherichia coli cell division amidase AmiA. We show that AmiA is a zinc metalloprotease that requires at least a tetrasaccharide glycopeptide substrate for cleavage. The approach outlined here can be applied to many other cell wall hydrolases and should enable more detailed studies of accessory proteins proposed to regulate amidase activity in cells.
三种周质 N-乙酰胞壁酰-L-丙氨酸酰胺酶对于水解肽聚糖的隔室至关重要，这使得细胞能够分离。酰胺酶裂解乳酰基和 L-丙氨酸的氨基之间的酰胺键，释放出肽部分。由于缺乏适合研究的底物，细胞分裂酰胺酶在很大程度上仍未被充分描述。在这里，我们使用了具有明确定义组成的合成肽聚糖片段来表征重要的大肠杆菌细胞分裂酰胺酶 AmiA 的催化活性和底物特异性。我们表明，AmiA 是一种锌金属蛋白酶，至少需要四糖糖肽底物才能进行切割。这里概述的方法可以应用于许多其他细胞壁水解酶，并且应该能够更详细地研究被提议调节细胞中酰胺酶活性的辅助蛋白。
