ABSTRACT
Circular dichroism (CD) spectroscopy is a well-established biophysical technique used to investigate the structure of molecules. The analysis of a protein CD spectrum depends on the quality of the original CD data, which can be affected by the sample purity, background absorption of the additives/solvent/buffer, the choice of the parameters used for data collection, etc. In this paper, the CD spectrum of myoglobin was used as a model to exploit how variations on each data collection parameter could affect the final protein CD spectrum and, the subsequent effect of them on the quantitative analysis of protein secondary structure. Bioinformatics analysis carried out with SESCA package and PDBMD2CD server predicted a theoretical myoglobin CD spectrum, and a Monte Carlo-like model was implemented to estimate the uncertainty in secondary structure predictions performed with CDSSTR, Selcon 3 and ContinLL algorithms. An inappropriate choice of data collection parameters can lead to a misinterpretation of the CD data in terms of the protein structural content.
Subject(s)
Circular Dichroism , Data Collection , Myoglobin , Protein Structure, Secondary , Reproducibility of ResultsABSTRACT
Antimicrobial peptides are a large group of natural compounds which present promising properties for the pharmaceutical and food industries, such as broad-spectrum activity, potential for use as natural preservatives, and reduced propensity for development of bacterial resistance. Plantaricin 149 (Pln149), isolated from Lactobacillus plantarum NRIC 149, is an intrinsically disordered peptide with the ability to inhibit bacteria from the Listeria and Staphylococcus genera, and which is capable of promoting inhibition and disruption of yeast cells. In this study, the interactions of Pln149 with model membranes composed of zwitterionic and/or anionic phospholipids were investigated using a range of biophysical techniques, including isothermal titration calorimetry, surface tension measurements, synchrotron radiation circular dichroism spectroscopy, oriented circular dichroism spectroscopy, and optical microscopy, to elucidate these peptides' mode of interactions and provide insight into their functional roles. In anionic model membranes, the binding of Pln149 to lipid bilayers is an endothermic process and induces a helical secondary structure in the peptide. The helices bind parallel to the surfaces of lipid bilayers and can promote vesicle disruption, depending on peptide concentration. Although Pln149 has relatively low affinity for zwitterionic liposomes, it is able to adsorb at their lipid interfaces, disturbing the lipid packing, assuming a similar parallel helix structure with a surface-bound orientation, and promoting an increase in the membrane surface area. Such findings can explain the intriguing inhibitory action of Pln149 in yeast cells whose cell membranes have a significant zwitterionic lipid composition.
