S-layers commonly cover archaeal cell envelopes and are composed of proteins that
self-assemble into a paracrystalline surface structure. Despite their detection
in almost all archaea, there are few reports investigating the structural
properties of these proteins, with no reports exploring this topic for
halophilic S-layers. The objective of the present study was to investigate the
secondary and tertiary organization of the
Haloferax volcanii
S-layer protein. Such investigations were performed using circular dichroism,
fluorescence spectroscopy, dynamic light scattering and transmission electron
microscopy. The protein secondary structure is centered on β-sheets and is
affected by environmental pH, with higher disorder in more alkaline conditions.
The pH can also affect the protein’s tertiary structure, with higher tryptophan
side-chain exposure to the medium under the same conditions. The concentrations
of Na, Mg and Ca ions in the environment also affect the protein structures,
with small changes in α-helix and β-sheet content, as well as changes in
tryptophan side chain exposure. These changes in turn influence the protein’s
functional properties, with cell envelope preparations revealing striking
differences when in different salt conditions. Thermal denaturation assays
revealed that the protein is stable. It has been reported that the S-layer
protein
N
-glycosylation process is affected by external factors
and the present study indicates for the first time changes in the protein
structure.
Background: visceral leishmaniasis (VL) is a critical public health problem in over ninety countries. The control measures adopted in Brazil have been insufficient when it comes to preventing the spread of this overlooked disease. In this context, a precise diagnosis of VL in dogs and humans could help to reduce the number of cases of this disease. Distinct studies for the diagnosis of VL have used single recombinant proteins in serological assays; however, the results have been variable, mainly in relation to the sensitivity of the antigens. In this context, the development of multiepitope-based proteins could be relevant to solving such problem. Methods: a chimeric protein (rMELEISH) was constructed based on amino acid sequences from kinesin 39 (k39), alpha-tubulin, and heat-shock proteins HSP70 and HSP 83.1, and tested in enzyme-linked immunosorbent (ELISA) for the detection of L. infantum infection using canine (n = 140) and human (n = 145) sera samples. Results: in the trials, rMELEISH was able to discriminate between VL cases and cross-reactive diseases and healthy samples, with sensitivity and specificity values of 100%, as compared to the use of a soluble Leishmania antigenic extract (SLA). Conclusions: the preliminary data suggest that rMELEISH has the potential to be tested in future studies against a larger serological panel and in field conditions for the diagnosis of canine and human VL.
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