Case study 2: Modulation excitation spectroscopy (MES)

Abstract

In this case of study, we provide examples of the use of infrared spectroscopy in attenuated total reflection (ATR) to investigate enzymes at work. An optimized ATR flow-thought cell was developed, which allows performing modulation excitation spectroscopy with phase sensitive detection (MES-PSD) experiments in a controlled way. The reversible effect of solvents with different hydrophobicity on the secondary structure of a Thermomyces lanuginosus lipase (TTL) was demonstrated by an analysis of the Amide I band. Finally, the lipase B of Candida antarctica (CALB), active to the enantioselective esterification of ketoprofen with alcohols was studied. MES-PSD analysis allowed to univocally identifying signals belonging to the interaction between ketoprofen and the lipase, that is the acyl enzyme intermediate.