Expression of the alpha subunit of PABA peptide hydrolase (EC 3.4.24.18) in MDCK cells Synthesis and secretion of an enzymatically inactive homodimer

AbstractIn this paper, we report the expression of PPHα in the polarized cell line MDCK (Madin Darby canine kidney). In these cells, the enzyme was synthesized m an inactive profonn, which upon treatment with trypsin was activated. The enzyme isolated from cell extracts was core-glycosylated and appeared to be retained in the ER as a homodimer. No PPHα was detectable on the surface of intact cells by immunofluoreseence. However, a complex glycosylated soluble but inactive form was present in the culture medium, suggesting that proteolytic removal of the C-terminal membrane anchoring peptide leads to the secretion of PPHα

Kullback-Leibler and Renormalized Entropy: Applications to EEGs of Epilepsy Patients

Recently, renormalized entropy was proposed as a novel measure of relative entropy (P. Saparin et al., Chaos, Solitons & Fractals 4, 1907 (1994)) and applied to several physiological time sequences, including EEGs of patients with epilepsy. We show here that this measure is just a modified Kullback-Leibler (K-L) relative entropy, and it gives similar numerical results to the standard K-L entropy. The latter better distinguishes frequency contents of e.g. seizure and background EEGs than renormalized entropy. We thus propose that renormalized entropy might not be as useful as claimed by its proponents. In passing we also make some critical remarks about the implementation of these methods.Comment: 15 pages, 4 Postscript figures. Submitted to Phys. Rev. E, 199

Genetic architecture of EEG power spectra in early life

We measured the electroencephalogram (EEG) in 209 5 year old monozygotic (MZ) and dizygotic (DZ) twin pairs to estimate the relative contribution of genetic and environmental factors to EEG power spectra in early life. Data from same-sex and from opposite-sex twin pairs were used to test for sex differences in genetic influences. Results showed high concordance for EEGs of MZ twins for absolute and relative power in δ, θ, α1, α2, β1 and β2 bands. A model with additive genetic and unique environmental influences explained individual differences in both absolute and relative power in almost all bands and all electrode positions. Heritability of EEG power spectra was high. For absolute power the highest heritabilities were observed in θ, α1, α2 and β1 power bands (mean heritability 81, 81, 78, and 73%, respectively). Somewhat lower heritabilities were found in δ and β2 bands (mean heritability 55 and 64%, respectively). For relative power heritabilities were 63, 76, 71, 72, 68, and 65 for δ, θ, α1, α2, β1, and β2, respectively. Virtually no sex differences in heritability were found. These findings indicate that the background EEG is one of the most heritable characteristics in early life

The Metalloprotease Meprinβ Processes E-Cadherin and Weakens Intercellular Adhesion

BACKGROUND: Meprin (EC 3.4.24.18), an astacin-like metalloprotease, is expressed in the epithelium of the intestine and kidney tubules and has been related to cancer, but the mechanistic links are unknown. METHODOLOGY/PRINCIPAL FINDINGS: We used MDCK and Caco-2 cells stably transfected with meprin alpha and or meprin beta to establish models of renal and intestinal epithelial cells expressing this protease at physiological levels. In both models E-cadherin was cleaved, producing a cell-associated 97-kDa E-cadherin fragment, which was enhanced upon activation of the meprin zymogen and reduced in the presence of a meprin inhibitor. The cleavage site was localized in the extracellular domain adjacent to the plasma membrane. In vitro assays with purified components showed that the 97-kDa fragment was specifically generated by meprin beta, but not by ADAM-10 or MMP-7. Concomitantly with E-cadherin cleavage and degradation of the E-cadherin cytoplasmic tail, the plaque proteins beta-catenin and plakoglobin were processed by an intracellular protease, whereas alpha-catenin, which does not bind directly to E-cadherin, remained intact. Using confocal microscopy, we observed a partial colocalization of meprin beta and E-cadherin at lateral membranes of incompletely polarized cells at preconfluent or early confluent stages. Meprin beta-expressing cells displayed a reduced strength of cell-cell contacts and a significantly lower tendency to form multicellular aggregates. CONCLUSIONS/SIGNIFICANCE: By identifying E-cadherin as a substrate for meprin beta in a cellular context, this study reveals a novel biological role of this protease in epithelial cells. Our results suggest a crucial role for meprin beta in the control of adhesiveness via cleavage of E-cadherin with potential implications in a wide range of biological processes including epithelial barrier function and cancer progression

Brown Spider (Loxosceles genus) Venom Toxins: Tools for Biological Purposes

Venomous animals use their venoms as tools for defense or predation. These venoms are complex mixtures, mainly enriched of proteic toxins or peptides with several, and different, biological activities. In general, spider venom is rich in biologically active molecules that are useful in experimental protocols for pharmacology, biochemistry, cell biology and immunology, as well as putative tools for biotechnology and industries. Spider venoms have recently garnered much attention from several research groups worldwide. Brown spider (Loxosceles genus) venom is enriched in low molecular mass proteins (5–40 kDa). Although their venom is produced in minute volumes (a few microliters), and contain only tens of micrograms of protein, the use of techniques based on molecular biology and proteomic analysis has afforded rational projects in the area and permitted the discovery and identification of a great number of novel toxins. The brown spider phospholipase-D family is undoubtedly the most investigated and characterized, although other important toxins, such as low molecular mass insecticidal peptides, metalloproteases and hyaluronidases have also been identified and featured in literature. The molecular pathways of the action of these toxins have been reported and brought new insights in the field of biotechnology. Herein, we shall see how recent reports describing discoveries in the area of brown spider venom have expanded biotechnological uses of molecules identified in these venoms, with special emphasis on the construction of a cDNA library for venom glands, transcriptome analysis, proteomic projects, recombinant expression of different proteic toxins, and finally structural descriptions based on crystallography of toxins