Metal binding to the surface lipoprotein AdcAI from Streptococcus pyogenes and to the salivary antimicrobial peptide Histatin-5

Abstract

The family of human salivary His-rich antimicrobial peptide, histatins, as exemplified by Histatin-5 (Hst5), bind zinc (Zn) and copper (Cu), but the role of metal binding in histatins have not been fully elucidated. Whether Hst5 is involved in nutritional immunity by limiting Zn availability to cause bacterial Zn starvation or by increasing Cu availability to cause Cu toxicity was examined in this study. Streptococcus pyogenes or Group A Streptococcus (GAS) was used as a model organism as it represents the oral streptococci that make up approximately 60 % of the oral microbiome. The metal binding affinity of Hst5 was measured by equilibrium competition assays using colourimetric probes and was compared against the high-affinity Zn uptake protein AdcAI from GAS. These experiments confirmed that Hst5 binds Zn weakly, and it does not strongly influence Zn availability. In addition, Hst5 cannot compete for Zn binding with AdcAI, which has an affinity that is 8 orders of magnitude higher than that of Hst5. In contrast, Hst5 has a high affinity for Cu and therefore influences the Cu availability. However, it does not promote Cu toxicity in GAS by delivering excessive Cu into the cytoplasm. It rather protects from the accumulation of intracellular Cu in ΔcopA mutant strain that is hypersensitive to increased Cu availability due to the loss of ability to export intracellular Cu. Whether Cu induces Cu starvation in Cu-requiring microbes remains unsolved, but this study has opened the possibility of Hst5 providing a buffering effect from metal toxicity to maintain a healthy oral microbiome.