Heat-Treated Lysozyme Hydrochloride: A Study on Its Structural Modifications and Anti-SARS-CoV-2 Activity

Abstract

Lysozyme (E.C. 3.2.1.17), an about 14 kDa protein and pI 11, widely spread in nature, is present in humans mainly in milk, saliva, and intestinal mucus as a part of innate defense mechanisms. It is endowed with antimicrobial activity due to its action as an N-acetylmuramidase, cleaving the 1-4β glycosidic linkage in the peptidoglycan layer of Gram-positive bacteria. This antimicrobial activity is exerted only against a limited number of Gram-negative bacteria. Different action mechanisms are proposed to explain its activity against Gram-negative bacteria, viruses, and fungi. The antiviral activity prompted the study of a possible application of lysozyme in the treatment of SARS-CoV-2 infections. Among the different sources of lysozyme, the chicken egg albumen was chosen, being the richest source of this protein (c-type lysozyme, 129 amino acids). Interestingly, the activity of lysozyme hydrochloride against SARS-CoV-2 was related to the heating (to about 100 °C) of this molecule. A chemical–physical characterization was required to investigate the possible modifications of native lysozyme hydrochloride by heat treatment. The FTIR analysis of the two preparations of lysozyme hydrochloride showed appreciable differences in the secondary structure of the two protein chains. HPLC and NMR analyses, as well as the enzymatic activity determination, did not show significant modifications.

Lysozyme: A Natural Product with Multiple and Useful Antiviral Properties

“…However, the only study documenting the activity of lysozyme against SARS-CoV-2, using lysozyme from hen egg white, shows a significant activity when lysozyme is brought into direct contact with the virus at concentrations of 0.19 to 1.00 mg/mL, which are not toxic to Vero cells, and when the lysozyme-virus mixture is applied to the Vero cells at a second moment (the authors refer to this as pre-treatment and post-treatment, respectively). A clear antiviral activity is observed at the highest concentrations of 0.75 or 1.00 mg/mL, and it is clearly recognizable with the combination of pre-treatment + post-treatment and somewhat lower with the pre-treatment or post-treatment alone [81]. However, for the sake of clarity, it must be mentioned that the studies are at a preliminary stage, and there is still a great chance of receiving further insights.…”

“…Strong antiviral effect at dosages lower than those of the single drug [63] Heat-treated lysozyme against SARS-CoV-2 in vitro Highest activity when lysozyme was pre-incubated with the virus and re-added to already-infected cells [81]…”