Characterization of heterologously expressed fibril filaments, a shape and motility determining cytoskeletal protein of the helical bacterium Spiroplasma

Fibril is a constitutive filament forming cytoskeletal protein of unidentified fold, exclusive to members of genus Spiroplasma. It is hypothesized to undergo conformational changes necessary to bring about Spiroplasma motility through changes in body helicity. However, in the absence of a cofactor such as nucleotide that binds to the protein and drives polymerization, the mechanism driving conformational changes in fibril remains unknown. Sodium dodecyl sulphate (SDS) solubilized the fibril filaments and facilitated fibril purification by affinity chromatography. An alternate protocol for obtaining enriched insoluble fibril filaments has been standardized using density gradient centrifugation method. Visualization of purified protein using electron microscopy demonstrated that it forms filament bundles. Probable domain boundaries of fibril protein were identified based on mass spectrometric analysis of proteolytic fragments. Presence of both α-helical and β-sheet signatures in FT-IR measurements suggests that fibril filaments consist of assembly of folded globular domains, and not a β-strand based aggregation similar to amyloid fibrils.

The Increased RNase Activity of IRE1α in PBMCs from Patients with Rheumatoid Arthritis

Purpose: Despite recent advances in the diagnosis and treatment of rheumatoid arthritis (RA), this inflammatory disease remains a challenge to patients and physicians. Recent evidence highlights the contribution of endoplasmic reticulum (ER) stress in the pathogenesis and treatment of RA. Herein, we study the expression of the ER stress sensor inositol-requiring enzyme 1α (IRE1α), as well as XBP1 splicing and the regulated IRE1-dependent decay (RIDD), in peripheral blood mononuclear cells (PBMCs) from patients with RA compared with healthy controls. Methods: The PBMCs from blood samples of RA patients and healthy volunteers were isolated by a density gradient centrifugation method using Ficoll. The gene expression levels of GRP78/ Bip, IRE1, XBP1s, micro-RNAs (miRNAs) were evaluated by real-time PCR. Results: The expression of GRP78, IRE1, and XBP1s were increased in PBMCs of RA patients compared with healthy controls. We further show that the RIDD targets (miRNA-17, -34a, -96, and -125b) were downregulated in RA samples. Conclusion: This study can expand our knowledge on the importance of RNase activity of IRE1α in RA and may offer new potentials for developing novel diagnostic and/or therapeutic biomarkers.

Keberhasilan inseminasi buatan menggunakan semen sexing beku pada Sapi Persilangan Ongole

<p>The Artificial Insemination (AI) results in Beef cattle are expected to be male calf because they can increaseweight gain rapidly, so in this study AI was carried out by using sexing semen with Percoll density gradient centrifugation method containing Y sperm.This study was conducted to evaluate the success rate of AI using frozen sexing semen on Ongole Cross Breed. The study method used wasexperimental, comparing AI frozen sexing semen and frozen semen in 54 Ongole Cross Breed. The study variableswere Non Return Rate₁ (NRR₁), Non Return Rate₂ (NRR₂) and Conception Rate (CR).The study showed that the success rate of based on the value of NRR₁ and NRR₂AI using frozen sexing semen on Ongole Cross Breed is lower than un sexed frozen semen with the NRR₁ value is 73,91%; 93,54% while the value of NRR₂ was 69,56%; 83,87%. Based on CR value of frozen sexing semen higher than un sexed frozen semen showed CR valuewas 43,47%; 35,48%.</p>

The separation of clay mineral fractions with linear heavy liquid density gradient columns

Because of increased interest in mineralogical analysis of soils a rapid, generally applicable method to separate clay minerals is needed, and such a method is described here. The technique is a modification of the heavy liquid density gradient centrifugation method which is commonly applied in mineralogical practice. Modifications are:(1)The use of suitable surface active agents to overcome the flocculation problem.(2)The use of the ultra centrifuge at about 33,000 g to hasten sedimentation.(3)The use of a linear density gradient which supplies in addition a quick identification of the mineral composition.Preliminary experiments, evaluating different variables (e.g. cation form, ultrasonic treatment, influence of surfactant), reveal the scope and limitations of the procedure presented.Clay minerals of different density can be separated. However, if the clay minerals of a mixture have overlapping densities, or if they contain a series of mixed-layer minerals, only a broad fractionation is possible. This is shown by X-ray diffraction and electron microscope studies of isolated zones (or parts of them) which develop in the centrifuged density gradient columns containing various pre-treated natural clays or clay mixtures.