Abstract
PsaA (pneumococcal surface antigen A) is a S. pneumoniae virulence factor that belongs to the metal transport system. The Manganese PsaA binding has been associated with oxidative stress resistance becoming a pivotal element in the bacteria virulence. It has been shown that Zinc inhibits the Manganese acquisition and promotes bacteria toxicity. We have performed a PsaA conformational analysis both in the presence (Zn-rPsaA) and in the absence of Zinc (free-rPsaA). We performed experiments in the presence of different Zinc concentrations to determine the metal minimum concentration which induced a conformational change. The protein in free and Zn-binding condition was also studied in pH ranging 2.6-8.0 and in temperature ranging 25oC-85oC. pH experiments showed a decrease of fluorescence intensity only in acidic medium. Analysis of the heat-induced denaturation demonstrated that Zinc-binding promoted an increase in melting temperature from 55oC (free-rPsaA) to 78.8oC (Zn-rPsaA) according to fluorescence measurements. In addition, the rPsaA stabilization by Zinc was verified through analysis of urea and guanidine hydrochloride denaturation. Data showed that Zinc promoted an increase in the rPsaA stability and its removal by EDTA can lead to a PsaA intermediate conformation. These findings can be considered in the development of vaccines containing PsaA as antigen.
Highlights
The Streptococcus pneumoniae is a grampositive coccus, known as pneumococcus, the most common human respiratory pathogen responsible for the annual death of millions of children worldwide (Kadioglu et al 2008)
Our results showed an rPsaA conformational change upon Zinc binding characterized by an increased stability of the protein and the structural consequences of the zinc removal
In order to evaluate the role of Zinc on the structural stability of rPsaA, this protein was expressed and purified (Larentis et al 2011)
Summary
The Streptococcus pneumoniae is a grampositive coccus, known as pneumococcus, the most common human respiratory pathogen responsible for the annual death of millions of children worldwide (Kadioglu et al 2008). Pneumococcal surface antigen A (PsaA) is a virulent factor, a 37 kDa lipoprotein which is hydrophobic, immunogenic and genetically conserved in S. pneumonia serotypes (Larentis et al 2012). Pneumococci possess an ATP-binding cassette (ABC) transporter cation permease encoded by the psaBCA locus (Dintilhac and Claverys 1997). This transporter is composed of the products of three genes, two units of an ATP-binding protein (PsaB) and two units of a hydrophobic membrane protein (PsaC), and a solute-binding lipoprotein (PsaA) organized in an operon with a gene encoding PsaD, a thiol peroxidase (Novak et al 1998). The β strands within each domain form parallel sheets and a metal-binding site is formed in the domain interface by the side chains of His, His139, Glu205 and Asp280 (Lawrence et al 1998)
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