News Release

Understanding variations in Salmonella virulence

University of Seville research provides insight into why Salmonella enzymes have certain specific human substrates and are not broad-spectrum

Peer-Reviewed Publication

University of Seville

Researchers from the University of Seville, in collaboration with the universities of Zaragoza and Kansas (USA), have managed to describe hitherto unknown processes that allow us to understand the virulence of a Salmonella infection. Specifically, their study, recently published in the journal Chemical Science, has shown that, in certain Salmonella virulence factors, a single amino acid is responsible for determining which proteins are modified in the infected cell.

Escherichia coli and Salmonella enterica are enterobacteria responsible for a multitude of cases of foodborne infections, typically associated with diarrhoea, fever, and nausea, with varying degrees of severity. Over the course of their evolution, these microbes have developed a whole strategic arsenal that they use during the infection process to be able to resist our cells’ natural defence mechanisms, as well as to promote their dissemination in the infected tissues (microbial invasion). An important part of this arsenal are the so-called ‘virulence factors’, which are molecules, typically proteins, that these microbes use to weaken the infected cells’ natural immune response towards the invader, thus aiding its dissemination in the infected organism.

These virulence factors include the bacterial enzymes themselves, which induce chemical modifications in infected cells, facilitating infection and subsequent microbial invasion. For these modifications to occur in the cell, one important aspect is that these enzymes must bind to certain proteins of the infected cell. Thus, a very promising way to prevent microbial invasion is to inhibit this interaction, that is, to prevent these molecules, from the invader and the infected cell, from binding together. This can be achieved by using molecules designed to occupy the site of the bacterial enzyme to which the cellular protein binds. However, for the design of these molecules (called inhibitors) to be effective, it is essential to know these enzymes’ three-dimensional characteristics in detail, as well as to know which of these enzymes’ fundamental components, i.e. amino acids, are responsible for the enzymatic activity that gives them virulence.

“Our interest was in analysing which amino acids in certain Salmonella virulence factors were responsible for making these enzymes more selective, i.e. modifying fewer proteins in the infected cell compared to homologous enzymes in Escherichia coli. We focused on enzymes called glycosyltransferases, which are a central theme in our laboratory, and we observed, through activity studies with different mutants that, just by changing an amino acid, that is, a simple mutation, these Salmonella enzymes recovered the ability to affect a greater number of substrates and thus show an activity similar to the equivalent enzymes of Escherichia coli,” explains Ramón Hurtado-Guerrero, professor at the University of Zaragoza.

The study is of great interest, since it not only opens the door to understanding the molecular basis of why Salmonella enzymes have certain specific human substrates and are not broad-spectrum like the enzyme in E. coli. Rather, the study has also made it possible to determine how the pathogenicity of Salmonella enterica can be varied with just simple changes in these enzymes, making it less or more pathogenic depending on the mutation.

“An important aspect was to elucidate the impact that the single mutation has on the binding between the virulence factor and the protein of the infected cell,” explains Jesús Angulo, a researcher at the University of Seville. “In our case, we studied why the single mutation of the enzyme called SseK1 in Salmonella makes it active in modifying a particular protein, called FADD, found in the infected cell, which it would not modify under ‘normal’ conditions. To do this, we carried out molecular dynamics calculations that showed that this mutation, which is not in the ‘active site’ of the enzyme, makes the enzyme and the substrate protein bind more tightly, since the mutated amino acid improves the fit between the contact surfaces of both molecules (akin to improving the contour of a key so that it fits the lock better). Furthermore, these molecular dynamics simulations showed that the single mutation favourably affected the dynamics of the catalytic site, such as to favour the chemical modification of the infected cell’s host protein. It is a very striking example of how a simple mutation can broaden the spectrum of action of an enzyme by simultaneously improving the binding affinity and the dynamics of the residues involved in the catalytic process,” explains Jesús Angulo.

These results are of great interest in the molecular understanding of the action of bacterial virulence factors, and may inspire the development of inhibitors as alternative antibacterial therapies to antibiotics.

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