Investigation of the role of the disulphide bond in the activity and structure of staphylococcal enterotoxin C1

126

We describe the complete sequence of the 15.9-kb staphylococcal pathogenicity island 3 encoding staphylococcal enterotoxin serotypes B, K, and Q. The island, which meets the generally accepted definition of pathogenicity islands, contains 24 open reading frames potentially encoding proteins of more than 50 amino acids, including an apparently functional integrase. The element is bordered by two 17-bp direct repeats identical to those found flanking staphylococcal pathogenicity island 1. The island has extensive regions of homology to previously described pathogenicity islands, particularly staphylococcal pathogenicity islands 1 and bov. The expression of 22 of the 24 open reading frames contained on staphylococcal pathogenicity island 3 was detected either in vitro during growth in a laboratory medium or serum or in vivo in a rabbit model of toxic shock syndrome using DNA microarrays. The effect of oxygen tension on staphylococcal pathogenicity island 3 gene expression was also examined. By comparison with the known staphylococcal pathogenicity islands in the context of gene expression described here, we propose a model of pathogenicity island origin and evolution involving specialized transduction events and addition, deletion, or recombination of pathogenicity island "modules."

Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 99%

Characterization and Expression Analysis of Staphylococcus aureus Pathogenicity Island 3

Yarwood

McCormick

Paustian

et al. 2002

126

“…Each animal received 1.0 ml of toxin solution per kg of body weight so that the dose received was 10 g/kg. We had determined previously that this dose of native SEC1 is sufficient to induce emesis in 100% of animals tested using this technique (9). Animals were returned to their cages following removal of the nasogastric tube and observed for 24 h.…”

Section: Methodsmentioning

confidence: 99%

“…Although TSS and staphylococcal food poisoning have overlapping symptoms, the pathogenesis of each is unique, and the biological activities responsible for both diseases are determined by separate regions of the SE molecules (7)(8)(9). The route of exposure also determines which disease will result.…”

mentioning

confidence: 99%

Zinc-mediated Dimerization and Its Effect on Activity and Conformation of Staphylococcal Enterotoxin Type C

Chi¹,

Sadler²,

Jabloński³

et al. 2002

Self Cite

Staphylococcal enterotoxins are superantigen exotoxins that mediate food poisoning and toxic shock syndrome in humans. Despite their structural and functional similarities, superantigens display subtle differences in biological properties and modes of receptor binding as a result of zinc atoms bound differently in their crystal structures. For example, the crystal structures of the staphylococcal enterotoxins in the type C serogroup (SECs) contain a zinc atom coordinated by one aspartate and two histidine residues from one molecule and another aspartate residue from the next molecule, thus forming a dimer. This type of zinc ligation and zinc-mediated dimerization occurs in several SECs, but not in most other staphylococcal enterotoxin serogroups. This prompted us to investigate the potential importance of zinc in SEC-mediated pathogenesis. Site-directed mutagenesis was used to replace SEC zinc binding ligands with alanine. SEC mutants unable to bind zinc did not have major conformational alterations although they failed to form dimers. Zinc binding was not essential for T cell stimulation, emesis, or lethality although in general the mutants were less pyrogenic. Thus the zinc atom in SECs might represent a non-functional heavy atom in an exotoxin group that has diverged from related bacterial toxins containing crucial zinc atoms.

“…Functional orthologs of Dsb proteins in Gram-positive organisms include Bacillus brevis Bdb (a DsbA ortholog) (16), Bacillus subtilis BdbB and BdbC (orthologs of DsbB), B. subtilis BdbD (a DsbA ortholog) (17)(18)(19)(20)(21) and Mycobacterium tuberculosis DsbE (structurally a homolog of DsbE, but functionally an oxidant like DsbA) (22). Moreover, functional virulence factors produced by Gram-positive bacteria, such as the human pathogen Staphylococcus aureus, contain disulfide bonds that are introduced during folding (23)(24)(25).…”

mentioning

confidence: 99%

Staphylococcus aureus DsbA Does Not Have a Destabilizing Disulfide

Heras

Kurz

Jarrott

et al. 2008

In Gram-negative bacteria, the introduction of disulfide bonds into folding proteins occurs in the periplasm and is catalyzed by donation of an energetically unstable disulfide from DsbA, which is subsequently re-oxidized through interaction with DsbB. Gram-positive bacteria lack a classic periplasm but nonetheless encode Dsb-like proteins. Staphylococcus aureus encodes just one Dsb protein, a DsbA, and no DsbB. Here we report the crystal structure of S. aureus DsbA (SaDsbA), which incorporates a thioredoxin fold with an inserted helical domain, like its Escherichia coli counterpart EcDsbA, but it lacks the characteristic hydrophobic patch and has a truncated binding groove near the active site. These findings suggest that SaDsbA has a different substrate specificity than EcDsbA. Thermodynamic studies indicate that the oxidized and reduced forms of SaDsbA are energetically equivalent, in contrast to the energetically unstable disulfide form of EcDsbA. Further, the partial complementation of EcDsbA by SaDsbA is independent of EcDsbB and biochemical assays show that SaDsbA does not interact with EcDsbB. The identical stabilities of oxidized and reduced SaDsbA may facilitate direct re-oxidation of the protein by extracellular oxidants, without the need for DsbB.The formation of native disulfide bonds through air oxidation of cysteine pairs is a slow reaction and organisms ranging from bacteria to humans encode enzymatic systems to catalyze the process. In eukaryotes, oxidative folding in the endoplasmic reticulum is primarily catalyzed by protein-disulfide isomerases that are reoxidized by Ero1p/Erv2p proteins (reviewed in Ref. 1). In Escherichia coli, dithiol oxidation takes place in the periplasm through the action of the Dsb 3 (Disulfide bond) family of proteins. Dsb proteins form two distinct pathways, the DsbA-DsbB (oxidative) pathway that introduces disulfides indiscriminately, and the DsbC/DsbG-DsbD (isomerization) pathway that shuffles incorrect disulfides (2, 3).Probably the best-studied Dsb protein is EcDsbA, the primary disulfide catalyst in E. coli (reviewed in Refs. 2, 4). This promiscuously oxidizing protein is a 21-kDa monomer containing a CPHC active site in a thioredoxin (TRX) fold with an inserted helical domain of ϳ80 residues. Upon catalyzing disulfide bond formation in substrate proteins, reduced EcDsbA relies on EcDsbB, a quinone reductase, to recover its catalytically active, and higher energy, oxidized form (5