Detection of Histatherin : a Potential Antimicrobial Peptide

Human health has advanced significantly since the discovery and development of antibiotics in the early 20 century. In recent times, due to a combination of overusing, misusing and declining research and development of antibiotics, resistance among pathogens has become endemic. Since the 1950’s antimicrobial peptides produced by higher order organisms such as cows, sheep and goats have been investigated. However studies into large-scale development of using antimicrobial peptides as a form of antibiotic have not become significant until recent years. One potential antimicrobial peptide identified in bovine milk and saliva is Histatherin. Studies into the synthetic version of Histatherin have shown antimicrobial activity against Staphylococcus aureus, Escherichia coli and Candida. The aim of this research was to identify the presence of the native Histatherin in bovine tissue samples, produce a purified fraction of Histatherin for antibody generation and to test the true antimicrobial activity of the native peptide as opposed to the synthetic. Initial studies were carried out using High Performance Liquid Chromatography (HPLC) to fractionate samples of lactoferrin eluate, 30 – 50 kDa permeate and bovine saliva. Western blots were conducted to detect the presence of Histatherin in these samples. Western blots conducted on lactoferrin eluate and bovine saliva fractions indicated positive results. However, larger starting volumes of lactoferrin eluate and saliva, using higher capacity columns may be needed to produce a more purified fraction of Histatherin. Due to some bands on the western blot being larger than 6 kDa in size it is thought that post-translational modifications or binding to other factors may be an explanation for this. Immunohistochemistry was finally carried out to identify the presence of Histatherin in bovine tissue samples, however the results are inconclusive at this stage.