Inntags: small self-structured epitopes for innocuous protein tagging.

Protein tagging is widely used in approaches ranging from affinity purification to fluorescence-based detection in live cells. However, an intrinsic limitation of tagging is that the native function of the protein may be compromised or even abolished by the presence of the tag. Here we describe and characterize a set of small, innocuous protein tags (inntags) that we anticipate will find application in a variety of biological techniques.

Antigen-Antibody Affinity for Dry Eye Biomarkers by Label Free Biosensing. Comparison with the ELISA Technique.

The specificity and affinity of antibody-antigen interactions is a fundamental way to achieve reliable biosensing responses. Different proteins involved with dry eye dysfunction: ANXA1, ANXA11, CST4, PRDX5, PLAA and S100A6; were validated as biomarkers. In this work several antibodies were tested for ANXA1, ANXA11 and PRDX5 to select the best candidates for each biomarker.

C-terminal truncation of the peroxiredoxin Tpx1 decreases its sensitivity for hydrogen peroxide without compromising its role in signal transduction.

Peroxiredoxins (Prxs) participate in hydrogen peroxide (H2O2) scavenging. Eukaryotic Prxs suffer H2O2-dependent inactivation, due to the oxidation of its catalytic cysteine to sulfinic acid, a modification which can be enzymatically reversed. This substrate-mediated reversible inactivation has been suggested to allow eukaryotic Prxs to act as floodgates, permitting high levels of H2O2 to trigger signal transduction.

The peroxiredoxin Tpx1 is essential as a H2O2 scavenger during aerobic growth in fission yeast.

Peroxiredoxins are known to interact with hydrogen peroxide (H(2)O(2)) and to participate in oxidant scavenging, redox signal transduction, and heat-shock responses. The two-cysteine peroxiredoxin Tpx1 of Schizosaccharomyces pombe has been characterized as the H(2)O(2) sensor that transduces the redox signal to the transcription factor Pap1. Here, we show that Tpx1 is essential for aerobic, but not anaerobic, growth.

Oxidative stress in Schizosaccharomyces pombe: different H2O2 levels, different response pathways.

Schizosaccharomyces pombe triggers different signalling pathways depending on the severity of the oxidative stress exerted, the main ones being the Pap1 and the Sty1 pathways. The Pap1 transcription factor is more sensitive to hydrogen peroxide (H(2)O(2)) than the MAP kinase Sty1 pathway, and is designed to induce adaptation, rather than survival, responses. The peroxiredoxin Tpx1 acts as a H(2)O(2) sensor and the upstream activator of the Pap1 pathway.

Differences in LexA regulon structure among Proteobacteria through in vivo assisted comparative genomics.

The LexA regulon encompasses an ensemble of genes involved in preserving cell viability under massive DNA damage and is present in most bacterial phyla. Up to date, however, the scope of this network had only been assessed in the Gamma Proteobacteria. Here, we report the structure of the LexA regulon in the Alpha Proteobacteria, using a combined approach that makes use of in vitro and in vivo techniques to assist and validate the comparative genomics in silico methodology.

Widespread distribution of a lexA-regulated DNA damage-inducible multiple gene cassette in the Proteobacteria phylum.

The SOS response comprises a set of cellular functions aimed at preserving bacterial cell viability in front of DNA injuries. The SOS network, negatively regulated by the LexA protein, is found in many bacterial species that have not suffered major reductions in their gene contents, but presents distinctly divergent LexA-binding sites across the Bacteria domain. In this article, we report the identification and characterization of an imported multiple gene cassette in the Gamma Proteobacterium Pseudomonas putida that encodes a LexA protein, an inhibitor of cell division (SulA), an error-prone polymerase (DinP) and the alpha subunit of DNA polymerase III (DnaE).

Geobacter sulfurreducens has two autoregulated lexA genes whose products do not bind the recA promoter: differing responses of lexA and recA to DNA damage.

The Escherichia coli LexA protein was used as a query sequence in TBLASTN searches to identify the lexA gene of the delta-proteobacterium Geobacter sulfurreducens from its genome sequence. The results of the search indicated that G. sulfurreducens has two independent lexA genes designated lexA1 and lexA2.

Role of the high-affinity zinc uptake znuABC system in Salmonella enterica serovar typhimurium virulence.

The Salmonella enterica serovar Typhimurium znuABC genes encoding a high-affinity zinc uptake system and its regulatory zur gene have been cloned. Salmonella serovar Typhimurium zur and znuC knockout mutants have been constructed by marker exchange. The 50% lethal dose of the znuC mutant increased when either orally or intraperitoneally inoculated in BALB/c mice, while virulence of the zur mutant decreased only when mice were intraperitoneally challenged.

Intracellular cyclic AMP concentration is decreased in Salmonella typhimurium fur mutants.

It is known that the Fur protein negatively regulates iron-uptake systems in different bacterial species, including Salmonella typhimurium. In this study it has been shown that the intracellular concentration of cyclic AMP (cAMP) is lower in a knockout S. typhimurium fur mutant than in the wild-type strain.