Gerstmann-Sträussler-Scheinker disease revisited: accumulation of covalently-linked multimers of internal prion protein fragments

Despite their phenotypic heterogeneity, most human prion diseases belong to two broadly defined groups: Creutzfeldt-Jakob disease (CJD) and Gerstmann-Sträussler-Scheinker disease (GSS). While the structural characteristics of the disease-related proteinase K-resistant prion protein (resPrPD) associated with the CJD group are fairly well established, many features of GSS-associated resPrPD are unclear. Electrophoretic profiles of resPrPD associated with GSS variants typically show 6-8 kDa bands corresponding to the internal PrP fragments as well as a variable number of higher molecular weight bands, the molecular nature of which has not been investigated. Here we have performed systematic studies of purified resPrPD species extracted from GSS cases with the A117V (GSSA117V) and F198S (GSSF198S) PrP gene mutations. The combined analysis based on epitope mapping, deglycosylation treatment and direct amino acid sequencing by mass spectrometry provided a conclusive evidence that high molecular weight resPrPD species seen in electrophoretic profiles represent covalently-linked multimers of the internal ~ 7 and ~ 8 kDa fragments. This finding reveals a mechanism of resPrPD aggregate formation that has not been previously established in prion diseases

Banding pattern indicative of echinococcosis in a commercial cysticercosis western blot

<p>Abstract</p> <p>Objective</p> <p>A commercial cysticercosis Western blot was evaluated for serological cross-reactivity of sera from patients with alveolar (AE) and cystic echinococcosis (CE).</p> <p>Methods</p> <p>A total of 161 sera were examined, including 31 sera from AE-patients, 11 sera from CE-patients, 9 sera from patients with other parasitic diseases and 109 sera from patients with unrelated medical conditions. All AE-and CE-sera were also examined by the echinococcosis Western blot.</p> <p>Results</p> <p>More sera from patients with AE than with CE showed cross-reactivity in the form of ladder-like patterns ("Mikado aspect") and untypical bands at 6-8 kDa (71% and 77.4% versus 27.3% and 45.5%, respectively). In contrast, triplets of bands in the area above 50 kDa and between 24 and 39-42 kDa were more frequent in CE than in AE sera. The fuzzy band at 50-55 kDa typical for cysticercosis was absent in all AE and CE sera.</p> <p>Conclusions</p> <p>Atypical banding patterns in the cysticercosis Western blot should raise the suspicion of a metacestode infection different from Taenia solium, i.e. Echinococcus multilocularis or E. granulosus, especially when the Mikado aspect and an altered 6-8 kDa band is visible in the absence of a fuzzy 50-55 kDa band.</p

Gerstmann-Sträussler-Scheinker disease subtypes efficiently transmit in bank voles as genuine prion diseases.

Gerstmann-Sträussler-Scheinker disease (GSS) is an inherited neurodegenerative disorder associated with mutations in the prion protein gene and accumulation of misfolded PrP with protease-resistant fragments (PrPres) of 6–8 kDa

Identification of a 68 kDa protein which copurifies with type-1 protein phosphatase as albumin

AbstractProteins of 60–70 kDa copurify with some preparations of type-1 or type-2 phosphatases. In our system chromatography on polylysine-Affi-Gel 10 separates a 68 kDa protein from rabbit muscle glycogen particle phosphorylase phosphatase. The separation affects neither the activity nor the size of the phosphatase. The 68 kDa protein, although pure by SDS gel electrophoresis criteria, still displays phosphatase activity of approx. 6–8 Umg. However, rechromatography either on Bio-Gel A-0.5 m or on Blue Sepharose CL-6B followed by gel filtration shows that the activity is due to a contamination with phosphatases of type 1 and type 2, displaying a molecular mass of 35 kDa, which can be totally removed from the 68 kDa protein. The amino acid composition of the 68 kDa protein is identical to that of rabbit serum albumin, within the limits of variation of the method. Furthermore, the sequence of the 38 N-terminal amino acids is the same in the isolated 68 kDa protein and in rabbit serum albumin

Death effector domain-containing protein induces vulnerability to cell cycle inhibition in triple-negative breast cancer

Lacking targetable molecular drivers, triple-negative breast cancer (TNBC) is the most clinically challenging subtype of breast cancer. In this study, we reveal that Death Effector Domain-containing DNA-binding protein (DEDD), which is overexpressed in > 60% of TNBCs, drives a mitogen-independent G1/S cell cycle transition through cytoplasm localization. The gain of cytosolic DEDD enhances cyclin D1 expression by interacting with heat shock 71 kDa protein 8 (HSC70). Concurrently, DEDD interacts with Rb family proteins and promotes their proteasome-mediated degradation. DEDD overexpression renders TNBCs vulnerable to cell cycle inhibition. Patients with TNBC have been excluded from CDK 4/6 inhibitor clinical trials due to the perceived high frequency of Rb-loss in TNBCs. Interestingly, our study demonstrated that, irrespective of Rb status, TNBCs with DEDD overexpression exhibit a DEDD-dependent vulnerability to combinatorial treatment with CDK4/6 inhibitor and EGFR inhibitor in vitro and in vivo. Thus, our study provided a rationale for the clinical application of CDK4/6 inhibitor combinatorial regimens for patients with TNBC

The electrophoretic membrane contactor: A mass-transfer-based methodology applied to the separation of whey proteins

In the electrophoretic membrane contactor (EMC), a porous membrane is used to establish a contact across two flowing liquids between which an electrically driven mass transfer takes places. In this work, a methodology is proposed to select the best operating conditions to separate biomolecules in an EMC. Single-solution experiments were coupled with a theoretical approach to predict the influence of the process parameters (pH, membrane MWCO) on the separation factor.This methodology was applied to the separation of wheyproteins, α-lactalbumin and β-lactoglobulin, which are known to be difficult to separate. Experiments were first carried out with single synthetic protein solutions at different pH values (4.8, 6 and 8) using cellulose acetate membranes of either 30 or 100 kDa molecular weight cut-off. The experimental work was associated with a theoretical approach to study the mass transfer mechanisms. The parameters used in the model were calculated from the experimental variations of the solute and solvent transfer. The dependence of these parameters on the operating conditions gives the extent of electrostatic repulsion and provides information on the steric effect with respect to separation performance.The model was then used to calculate the separation factor for various operating conditions in order to determine the best ones (pH and membrane) for fractionation. Using the results, fractions enriched in α-lactalbumin and in β-lactoglobulin were obtained at pH 4.8 with the 100 kDa membrane

Angiotensin I-converting enzyme inhibitor derived from cottonseed protein hydrolysate

Six proteolytic enzymes, including alcalase, flavourzyme, trypsin, neutrase, papain and pepsin, were employed to hydrolyze cottonseed protein to produce the hydrolysates of Angiotensin I-converting enzyme (ACE) inhibitory activity. The result indicated that the cottonseed protein hydrolysate (CPH) produced by papain had the highest ACE inhibitory activity. Therefore, papain was selected for enzymatic production of ACE inhibitor from cottonseed protein isolates (CPI). CPI was hydrolyzed withpapain for 1 - 8 h, and the 6 h hydrolysate had the strongest ACE inhibitory ability. The product was separated into four ranges of molecular weight (UF-I, &gt; 30 kDa; UF-II, 30 – 10 kDa; UF - III, 10 - 5 kDa; UF - IV, &lt; 5 kDa) by using an ultrafiltration (UF) membrane bioreactor system. Among them, UF-IV showed the highest ACE inhibitory activity (IC50 = 0.792 mg/ml). UF-IV was further fractionated with Sephadex G-25 gel filtration chromatography into four fractions (Fra I, Fra II, Fra III and Fra IV) that were composed of peptides of &gt;2.43 kDa, 2.43 - 0.82 kDa, 0.82 - 0.35 kDa and &lt;0.35 kDa, respectively. Fra II exhibited the strongest ACE inhibitory ability (IC50 = 0.159 mg/ml) with the yield of 41.63%. It was suggested that Fra II with good ACE inhibitory activity can be a potential source of natural ACE inhibitor.Keywords: Cottonseed protein hydrolysate, peptide fractions, angiotensin I-converting enzyme inhibitory ability, ultrafiltratio

A relevant IgE-reactive 28 kDa protein identified from Salsola kali pollen extract by proteomics is a natural degradation product of an integral 47 kDa polygalaturonase

[EN] A highly prevalent IgE-binding protein band of 28 kDa is observed when Salsola kali pollen extract is incubated with individual sera from Amaranthaceae pollen sensitized patients. By an immunoproteomic analysis of S. kali pollen extract, we identified this protein band as an allergenic polygalacturonase enzyme. The allergen, named Sal k 6, exhibits a pI of 7.14 and a molecular mass of 39,554.2 Da. It presents similarities to Platanaceae, Poaceae, and Cupressaceae allergenic polygalacturonases. cDNA-encoding sequence was subcloned into the pET41b vector and produced in bacteria as a His-tag fusion recombinant protein. The far-UV CD spectrum determined that rSal k 6 was folded. Immunostaining of the S. kali pollen protein extract with a rSal k 6-specific pAb and LC-MS/MS proteomic analyses confirmed the co-existence of the 28 kDa band together with an allergenic band of about 47 kDa in the pollen extract. Therefore, the 28 kDa was assigned as a natural degradation product of the 47 kDa integral polygalacturonase. The IgE-binding inhibition to S. kali pollen extract using rSal k 6 as inhibitor showed that signals directed to both protein bands of 28 and 47 kDa were completely abrogated. The average prevalence of rSal k 6 among the three populations analyzed was 30%, with values correlating well with the levels of grains/m(3) of Amaranthaceae pollen. Sal k 6 shares IgE epitopes with Oleaceae members (Fraxinus excelsior, Olea europaea and Syringa vulgaris), with IgE-inhibition values ranging from 20% to 60%, respectively. No IgE-inhibition was observed with plant-derived food extracts.This work was supported by grants SAF2011-26716 and SAF2014-53209-R from the Ministerio de Economia y Competitividad and RIRAAF Network RD12/0013/0015 from the ISCIII. R.B. was a fellow of the Ramon y Cajal program of the Ministerio de Economia y Competitividad (Spain). C.O-S. is supported by a contract of the Programa Operativo de Empleo Juvenil y la Iniciativa de Empleo Juvenil (YEI) with the participation of the Consejeria de Education, Juventud y Deporte de la Comunidad de Madrid y del Fondo Social Europeo.Mas-García, S.; Oeo-Santos, C.; Cuesta-Herranz, J.; Díaz-Perales, A.; Colás, C.; Fernández, J.; Barber, D.... (2017). A relevant IgE-reactive 28 kDa protein identified from Salsola kali pollen extract by proteomics is a natural degradation product of an integral 47 kDa polygalaturonase. Biochimica et Biophysica Acta (BBA) - Proteins and Proteomics. 1865(8):1067-1076. https://doi.org/10.1016/j.bbapap.2017.05.007S106710761865

Differential expression of collectins in human placenta and role in inflammation during spontaneous Labor.

© 2014 Yadav et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.Collectins, collagen-containing Ca2+ dependent C-type lectins and a class of secretory proteins including SP-A, SP-D and MBL, are integral to immunomodulation and innate immune defense. In the present study, we aimed to investigate their placental transcript synthesis, labor associated differential expression and localization at feto-maternal interface, and their functional implication in spontaneous labor. The study involved using feto-maternal interface (placental/decidual tissues) from two groups of healthy pregnant women at term (≥37 weeks of gestation), undergoing either elective C-section with no labor ('NLc' group, n = 5), or normal vaginal delivery with spontaneous labor ('SLv' group, n = 5). The immune function of SP-D, on term placental explants, was analyzed for cytokine profile using multiplexed cytokine array. SP-A, SP-D and MBL transcripts were observed in the term placenta. The 'SLv' group showed significant up-regulation of SP-D (p = 0.001), and down-regulation of SP-A (p = 0.005), transcripts and protein compared to the 'NLc' group. Significant increase in 43 kDa and 50 kDa SP-D forms in placental and decidual tissues was associated with the spontaneous labor (p<0.05). In addition, the MMP-9-cleaved form of SP-D (25 kDa) was significantly higher in the placentae of 'SLv' group compared to the 'NLc' group (p = 0.002). Labor associated cytokines IL-1α, IL-1β, IL-6, IL-8, IL-10, TNF-α and MCP-1 showed significant increase (p<0.05) in a dose dependent manner in the placental explants treated with nSP-D and rhSP-D. In conclusion, the study emphasizes that SP-A and SP-D proteins associate with the spontaneous labor and SP-D plausibly contributes to the pro-inflammatory immune milieu of feto-maternal tissues.Funding provided by BT/PR15227/BRB/10/906/2011) Department of Biotechnology (DBT), Government of India http://dbtindia.nic.in/index.asp (TM) and Indian Council of Medical Research (ICMR) Junior Research Fellowship (JRF)/Senior Research Fellowship (SRF), Government of India, www.icmr.nic.in (AKY)

Mobility of antimony, arsenic and lead at a former mine, Glendinning, Scotland

Elevated concentrations of antimony (Sb), arsenic (As) and lead (Pb) in upland organic-rich soils have resulted from past Sb mining activities at Glendinning, southern Scotland. Transfer of these elements into soil porewaters was linked to the production and leaching of dissolved organic matter and to leaching of spoil material. Sb was predominantly present in truly dissolved (&lt; 3 kDa) forms whilst As and Pb were more commonly associated with large Fe-rich/organic colloids. The distinctive porewater behaviour of Sb accounts for its loss from deeper sections of certain cores and its transport over greater distances down steeper sections of the catchment. Although Sb and As concentrations decreased with increasing distance down a steep gully from the main spoil heap, elevated concentrations (~ 6-8 and 13-20 μg L− 1, respectively) were detected in receiving streamwaters. Thus, only partial attenuation occurs in steeply sloping sections of mining-impacted upland organic-rich soils and so spoil-derived contamination of surface waters may continue over time periods of decades to centuries