scholarly journals The refolding activity of the yeast heat shock proteins Ssa1 and Ssa2 defines their role in protein translocation.

1996 ◽  
Vol 135 (5) ◽  
pp. 1229-1237 ◽  
Author(s):  
G L Bush ◽  
D I Meyer
Keyword(s):  
Protein Translocation ◽  
In Vitro Assay ◽  
Secretory Proteins ◽  
Vitro Assay ◽  
Chaperone Function ◽  
Cotranslational Folding ◽  
Yeast Lysate ◽  
Shock Proteins ◽  
First Time

Ssa1/2p, members of one of the yeast cytosolic hsp70 subfamilies, have been implicated in the translocation of secretory proteins into the lumen of the ER. The involvement of these hsp70s in translocation was tested directly by examining the effect of immunodepleting Ssa1/2p from yeast cytosol and subsequently testing the cytosol for its ability to support co- and post-translational translocation of prepro-alpha-factor. Depletion of Ssa1/2p had no effect on the efficiency of translocation in this in vitro assay. The system was used to examine the effect of the absence of Ssa1/2p on two other putative hsp70 functions: cotranslational folding of nascent luciferase and refolding of denatured luciferase. Depletion of Ssa1/2p had no effect on the ability of the yeast lysate to synthesize enzymatically active luciferase, but had a dramatic effect on the ability of the lysate to refold chemically denatured luciferase. These results demonstrate, for the first time, the refolding activity of Ssa1/2p in the context of the yeast cytosol, and define refolding activity as a chaperone function specific to Ssa1/2p, aprt from other cytosolic hsp70s. They also suggest that Ssa1/2p do not play a significant role in chaperoning the folding of nascent polypeptides. The implications of these findings for Ssa1/2p activity on their proposed role in the process of translocation are discussed.

scholarly journals Sac1p mediates the adenosine triphosphate transport into yeast endoplasmic reticulum that is required for protein translocation.

1995 ◽  
Vol 131 (6) ◽  
pp. 1377-1386 ◽  
Author(s):  
P Mayinger ◽  
V A Bankaitis ◽  
D I Meyer
Keyword(s):  
Endoplasmic Reticulum ◽  
Protein Translocation ◽  
Protein A ◽  
Lipid Vesicles ◽  
Secretory Proteins ◽  
Transport Activity ◽  
Vitro Assay ◽  
Microsomal Membranes

Protein translocation into the yeast endoplasmic reticulum requires the transport of ATP into the lumen of this organelle. Microsomal ATP transport activity was reconstituted into proteoliposomes to characterize and identify the transporter protein. A polypeptide was purified whose partial amino acid sequence demonstrated its identity to the product of the SAC1 gene. Accordingly, microsomal membranes isolated from strains harboring a deletion in the SAC1 gene (sac1 delta) were found to be deficient in ATP-transporting activity as well as severely compromised in their ability to translocate nascent prepro-alpha-factor and preprocarboxypeptidase Y. Proteins isolated from the microsomal membranes of a sac1 delta strain were incapable of stimulating ATP transport when reconstituted into the in vitro assay system. When immunopurified to homogeneity and incorporated into artificial lipid vesicles, Sac1p was shown to reconstitute ATP transport activity. Consistent with the requirement for ATP in the lumen of the ER to achieve the correct folding of secretory proteins, the sac1 delta strain was shown to have a severe defect in transport of procarboxypeptidase Y out of the ER and into the Golgi complex in vivo. The collective data indicate an intimate role for Sac1p in the transport of ATP into the ER lumen.

scholarly journals PTP1B INHIBITORS FROM ISODON TERNIFOLIUS COLLECTED IN VIETNAM

2020 ◽  
Vol 58 (5) ◽  
pp. 533
Author(s):  
Nguyen Phi-Hung
Keyword(s):  
Enzyme Activity ◽  
Inhibitory Activity ◽  
In Vitro Assay ◽  
Vitro Assay ◽  
Chemical Structures ◽  
Whole Plant ◽  
Ic50 Values ◽  
Ptp1b Inhibitors ◽  
First Time

From the whole plant of Isodon ternifolius collected in Vietnam, four triterpens including ursaldehyde (1), ursolic acid (2), b-sitosterol (3) and b-sitosteryl ferulate (4) were purified. Their chemical structures were determined by interpretation of NMR and MS data and comparison with the literatures. Compounds 1-4 were evaluated for their inhibitory activity against PTP1B enzyme activity using in vitro assay. Compounds 1 and 2 displayed potential activities with IC50 values of 16.92 ± 0.12 and 3.42 ± 0.45 μM, respectively. This is the first time that compounds 1 and 4 have been isolated from the Isodon genus and I. ternifolius has been evaluated for the PTP1B inhibitory activity.

scholarly journals Attachment Ligands of Viable Toxoplasma gondii Induce Soluble Immunosuppressive Factors in Human Monocytes

1999 ◽  
Vol 67 (5) ◽  
pp. 2547-2551 ◽  
Author(s):  
Jacqueline Y. Channon ◽  
Edward I. Suh ◽  
Rosanne M. Seguin ◽  
Lloyd H. Kasper
Keyword(s):  
Toxoplasma Gondii ◽  
Surface Antigen ◽  
Marked Decrease ◽  
In Vitro Assay ◽  
Surface Proteins ◽  
Secretory Proteins ◽  
Human Monocytes ◽  
Vitro Assay ◽  
Monocyte Cell

ABSTRACT Previous studies have demonstrated that surface antigen proteins, in particular SAG-1, of Toxoplasma gondii are important to this parasite as attachment ligands for the host cell. An in vitro assay was developed to test whether these ligands and other secretory proteins are involved in the immune response of human cells to toxoplasma. Human monocytes were infected with tachyzoites in the presence of antiparasite antibodies, and their effect on mitogen-induced lymphoproliferation was examined. The presence of antibody to either parasite-excreted proteins (MIC-1 and MIC-2) or surface proteins (SAG-1 and SAG-2) during infection neutralized the marked decrease seen in mitogen-induced lymphoproliferation in the presence of infected monocytes. Conversely, antibodies to other secreted proteins (ROP-1) and cytoplasmic molecules had no effect on parasite-induced, monocyte-mediated downregulation. Fluorescence microscope analysis detected microneme and surface antigen proteins on the monocyte cell surface during infection. These results suggest that microneme and surface antigen proteins trigger monocytes to downregulate mitogen-induced lymphoproliferation.

Vesicular Lipid Nanoparticles (Liposomes) for the Treatment of Medical Device Infections

2011 ◽  
Vol 1316 ◽  
Author(s):  
Erik Taylor ◽  
Anubhav Kaviratna ◽  
Rinti Banerjee ◽  
Thomas J. Webster
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Lipid Nanoparticles ◽  
In Vitro Assay ◽  
Silver Sulfadiazine ◽  
Human Pathogens ◽  
Vitro Assay ◽  
First Time ◽  
Better Than

AbstractLiposomes (a phospholipid bi-layer which can be formulated to contain drugs or other reagents) composed of endogenous phospholipid dipalmitoylphosphatidylcholine (DPPC) in combination with dioleoylphosphatidylethanolamine (DOPE), lauric acid, and silver sulfadiazine were made into vesicular nanoparticles in this study using an optimized extrusion technique. Liposomes were then tested for antibacterial activity against a range of bacteria species including Pseudomonas aeruginosa, Staphylococcus aureus, Escherichia coli, and Bacillus subtilis (all are relevant human pathogens known to infect implants) and were also challenged to prevent the growth of adherent biofilms (a robust slimy extracellular matrix) through an in vitro assay relevant to device related infections. It was found that all liposomes reduced bacterial growth, and, most importantly, liposomes containing DPPC and DOPE reduced biofilm formation better than the commercially available antibiotic silver sulfadiazine. These results indicated for the first time that such liposomes might be a better approach to prevent device related infections.

scholarly journals A new 11,10-guaiane-type sesquiterpenoid from the roots of Stellera chamaejasme Linn

2020 ◽  
pp. 174751982096104
Author(s):  
Fei-Feng Hu ◽  
Dan-Dan Qi ◽  
Sha Xu ◽  
Wei Mao
Keyword(s):  
Spectroscopic Analysis ◽  
Triple Negative ◽  
Cancer Cell Line ◽  
In Vitro Assay ◽  
Resonance Spectroscopy ◽  
Ionization Mass ◽  
Vitro Assay ◽  
Stellera Chamaejasme ◽  
First Time

Chamaejasmone F, a new 11,10-guaiane-type sesquiterpenoid, is isolated from the roots of Stellera chamaejasme Linn., along with four known compounds including two guaiane sesquiterpenoids and two lignans. The structure of the new compound is elucidated by extensive spectroscopic analysis, especially two-dimensional nuclear magnetic resonance spectroscopy and high-resolution electrospray ionization mass spectrometry. The known compounds 1β-hydroxy-10βH-guaia-4,11-dien-3-one and (+)-hinokinin are isolated from the genus Stellera for the first time. In an in vitro assay, chamaejasmone F shows moderate cytotoxic activity against the human triple negative breast cancer cell line MB-MDA-231.

In Vitro Assay of Platelet Adhesiveness with Washed and Tanned Human Red Cells

1968 ◽  
Vol 20 (03/04) ◽  
pp. 384-396 ◽  
Author(s):  
G Zbinden ◽  
S Tomlin
Keyword(s):  
Platelet Adhesion ◽  
Sodium Citrate ◽  
Blood Platelets ◽  
In Vitro Assay ◽  
Red Cells ◽  
Platelet Adhesiveness ◽  
Vitro Assay ◽  
In Vitro System ◽  
Human Red Cells

SummaryAn in vitro system is described in which adhesion of blood platelets to washed and tannic acid-treated red cells was assayed quantitatively by microscopic observation. ADP, epinephrine and TAME produced a reversible increase in platelet adhesiveness which was antagonized by AMP. With Evans blue, polyanetholsulfonate, phthalanilide NSC 38280, thrombin and heparin at concentrations above 1-4 u/ml the increase was irreversible. The ADP-induced increase in adhesiveness was inhibited by sodium citrate, EDTA, AMP, ATP and N-ethylmaleimide. EDTA, AMP and the SH-blocker N-ethylmaleimide also reduced spontaneous platelet adhesion to red cells. No significant effects were observed with adenosine, phenprocoumon, 5-HT, phthalanilide NSC 57155, various estrogens, progestogens and fatty acids, acetylsalicylic acid and similarly acting agents, hydroxylamine, glucose and KCN. The method may be useful for the screening of thrombogenic and antithrombotic properties of drugs.

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