Use of Atomic Force Microscopy as a Tool to Understand the Action of Antimicrobial Peptides on Bacteria
Atomic force microscopy (AFM) has been extensively used to image the three-dimensional surface morphology of a broad range of biological samples, including Gram-negative bacteria, imaged in the presence of antimicrobial peptides (AMPs). Although this technique provides high molecular resolution, it only requires minimum sample treatment and can even be performed in liquid and at varying temperatures while keeping the bacterial cells viable. In this chapter, we describe an easy, fast, and yet effective method for preparing AMP-treated Gram-negative bacteria samples for AFM imaging. The results obtained using this method show a series of morphological changes of Gram-negative bacteria upon treatment with selected AMPs, thus providing vivid insights into the mechanisms of how AMPs perturb and destroy Gram-negative bacteria in a stepwise manner. Technical details for performing AFM so as to obtain reliable and high-resolution images will also be discussed, together with some possible artifacts and troubleshooting.
Key wordsAtomic force microscopy Gram-negative bacteria bacteria surface morphology antimicrobial peptides lipopolysaccharide Sushi peptides
We thank Mr. Han Chong Ng for the preparation of the bacterial cultures and Ms. Pei Yun Lee for the assistance on AFM imaging and discussion. This work was funded by the Singapore-MIT Alliance (SMA2) and MoE Tier 2 grant (J.L. Ding and B. Ho).
- 6.Li, P., Wohland, T., Ho, B., and Ding, J. L. (2004) Perturbation of lipopolysaccharide (LPS) micelles by Sushi 3 (S3) antimicrobial peptide. The importance of an intermolecular disulfide bond in S3 dimer for binding, disruption, and neutralization of LPS. J. Biol. Chem. 279, 50150–50156.PubMedCrossRefGoogle Scholar