Structural basis for the interaction of the fluorescence probe 8-anilino-1-naphthalene sulfonate (ANS) with the antibiotic target MurA

摘要

The extrinsic fluorescence dye 8-anilino-1-naphthalene sulfonate (ANS) is widely used for probing conformational changes in pro- teins, yet no detailed structure of ANS bound to any protein has been reported so far.ANS has been successfully used to monitor the induced-fit mechanism of MurA [UDPGlcNAc enolpyruvyltrans- ferase (EC 2.5.1.7)], an essential enzyme for bacterial cell wall biosynthesis. We have solved the crystal structure of the ANS.MurA complex at 1.7-A resolution. ANS binds at an originally solvent. exposed region near Pro-112 and induces a major restructuring of the loop Pro-112-Pro-121 , such that a specific binding site emerges. The fluorescence probe is sandwiched between the strictly con- served residues Arg-91 . Pro-l12. and Gly-l13. Substrate binding to MurA is accompanied by large movements especially of the loop and Arg-91. which explains why ANS is an excellent sensor of conformational changes during catalysis of this pharmaceutically important enzyme.