The Adhesive Properties of MtsA in Group A Streptococcus

Abstract

Group A streptococcus (GAS) currently affects over half a million people a year worldwide. GAS is responsible for a wide range of diseases including the clinically uncomplicated pharyngitis and pyoderma but can lead to more serious and life-threatening conditions such as streptococcal toxic shock syndrome and necrotizing fasciitis. GAS harbours numerous virulence factors that are responsible for the pathophysiology of GAS. One group of these virulence factors is known as adhesins and is responsible for the attachment of the bacterium to the host cell. The only known host of GAS is the human whereby the skin and upper respiratory tract are the predominant sites of infection. These adhesins are surface exposed and include the fibronectin binding proteins and the M protein. Recently, there has been an increasing amount of evidence suggesting that bacterial proteins in metabolic pathways and house-keeping proteins may also play a role in adherence. The lipoprotein MtsA, is part of the GAS ATP binding cassette metal transporter which plays a role in the transportation of iron into the cell. In silico analysis revealed a 77% amino acid sequence identity with a Streptococcus pneumoniae adhesin known as pneumococcal surface adhesin A (PsaA). Recently, E-cadherin has been shown to be a possible receptor for PsaA. In this study, soluble and membrane-bound MtsA was generated and tested for the ability to bind to human keratinocytes and recombinant E-cadherin. The cloning vector pBlueScript was used to clone mtsA. The expression vector pET32a was used to express MtsA in E. coli whilst membrane-bound forms were generated by expressing MtsA in Lactococcus lactis (L. lactis). Antibodies were generated against MtsA using a New Zealand White rabbit which were subsequently used to detect the expression of MtsA in L. lactis by a Western blot. The antibodies were also used to detect MtsA adherence in enzyme-linked immunosorbent assays. No binding to human keratinocytes or E-cadherin was observed when recombinant MtsA was used. Likewise, no binding was observed using L. Lactis mutants expressing MtsA. However, flow cytometry revealed that MtsA was not exposed on the surface of the L. lactis mutant which could explain lack of binding observed when using L. lactis. Although it appears that MtsA does not act as an adhesin, with increasing evidence to suggest some bacterial proteins have more than one function, it may be that MtsA has a yet to be discovered alternate function.