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Antimicrobial peptides (AMPs) are generally cationic and amphipathic peptides that show potential applications to combat the growing threat of antibiotic resistant infections. AMPs are known to interact with bacterial membranes, but their mechanisms of toxicity and selectivity are poorly understood, in part because it is challenging to characterize AMP oligomeric complexes within lipid bilayers. Here, we used native mass spectrometry (MS) to measure the stoichiometry of AMPs inserted into lipoprotein nanodiscs with different lipid components. Titrations of increasing peptide concentration and collisional activation experiments reveal that AMPs can exhibit a range of behaviors from non-specific incorporation into the nanodisc to formation of specific complexes. This new approach to characterizing formation of AMP complexes within lipid membranes will provide unique insights into AMP mechanisms.

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Elucidation of fast chemical reactions such as protein folding requires resolution on a submillisecond time scale. However, most quench-flow and stop-flow techniques only allow chemical processes to be studied after a few milliseconds have elapsed. In order to shorten the minimum observation time for quench-flow experiments, we designed a miniaturized mixer assembly. Two “T” mixers connected by a channel are etched into a 1 cm × 1 cm silicon chip which is interfaced with a commercially available quench-flow instrument. Decreasing the volume of the mixing chambers and the distance between them results in an instrument with greatly reduced dead times. As a test of submillisecond measurements, we studied the basic hydrolysis of phenyl chloroacetate. This reaction proceeds with a second-order rate constant, k = 430 M-1 s-1, and shows pseudo-first-order kinetics at high hydroxide concentrations. The chemical reaction data demonstrate that the silicon device is capable of initiating and quenching chemical reactions in time intervals as short as 110 μs. The performance of these mixers was further confirmed by visualization using acid−base indicators.