scholarly journals The lanthanide-enhanced affinity chromatography of clostridial collagenase

1985 ◽  
Vol 225 (2) ◽  
pp. 553-556 ◽  
Author(s):  
C H Evans
Keyword(s):  
Periodic Acid ◽  
Sodium Dodecyl ◽  
Minor Component ◽  
Electrophoretic Analysis ◽  
Lanthanide Ions ◽  
Affinity Column ◽  
Major Protein ◽  
Periodic Acid Schiff ◽  
Highly Active ◽  
A Minor

Clostridiopeptidase A (EC 3.4.24.3) did not bind to a collagen affinity column in the absence of Ca2+, but did so in the presence of lanthanide ions (Ln3+). The sequestered enzyme could be eluted with EGTA. For the four Ln3+ ions tested, the order of efficiency in promoting enzyme binding, Sm3+ greater than Lu3+ greater than Er3+ much greater than La3+, reflected their relative abilities to inhibit clostridiopeptidase A. By using Sm3+ as an adjunct, it proved possible to separate a highly active preparation of collagenase from crude clostridial collagenase. Sodium dodecyl sulphate/polyacrylamide-gel-electrophoretic analysis of the preparation revealed a major protein of Mr 95000 and a minor component of Mr 82000. As both were stained by periodic acid/Schiff reagent, they were probably glycoproteins.

scholarly journals The burden of visceral leishmaniasis in Italy from 1982 to 2012: a retrospective analysis of the multi-annual epidemic that occurred from 1989 to 2009

2013 ◽  
Vol 18 (29) ◽  
Author(s):  
M Gramiccia ◽  
A Scalone ◽  
T Di Muccio ◽  
S Orsini ◽  
E Fiorentino ◽  
...  
Keyword(s):  
Visceral Leishmaniasis ◽  
Retrospective Analysis ◽  
Minor Component ◽  
Campania Region ◽  
Highly Active ◽  
Immunodeficiency Virus ◽  
Associated Conditions ◽  
A Minor ◽  
Immunocompetent Adults ◽  
Peak Value

Starting from 1989 Italy experienced an increase of visceral leishmaniasis (VL) cases over a baseline of 10 to 30 cases reported annually. The number of cases peaked in 2000 and 2004 with more than 200 cases/year, and subsequently declined to reach on average one third of the 2000 peak value in the period after 2010. A retrospective analysis from 1982 to 2012 showed that the multi-annual epidemic consisted of major components including (i) an outbreak involving infants and immunocompetent adults in parts of the Campania region (southern peninsular Italy) and that appears to have declined naturally, (ii) a second outbreak affecting human immunodeficiency virus (HIV)-infected individuals throughout the country, that declined owing to the use of highly active antiretroviral therapies (HAART), (iii) a generalised increase of VL cases in immunocompetent individuals and patients affected by associated conditions other than HIV from endemic regions of peninsular and insular Italy (other than Campania), which was due to a geographical spreading of VL foci, with no major case-clusters or outbreak features. A minor component consisted in the appearance of a few autochthonous cases in formerly non-endemic areas, starting from the early 1990s. Epidemic determinants and reasons for VL decline in the Campania region remain largely unexplained, despite the information available on canine reservoir and phlebotomine vectors in Italy.

The structure and associations of the double S layer on the cell wall of Aquaspirillum sinuosum

1990 ◽  
Vol 36 (5) ◽  
pp. 327-335 ◽  
Author(s):  
Stephen H. Smith ◽  
Robert G. E. Murray
Keyword(s):  
Cell Wall ◽  
Outer Membrane ◽  
Outer Layer ◽  
Peptide Mapping ◽  
Periodic Acid ◽  
Minor Component ◽  
Membrane Resistance ◽  
Surface Layers ◽  
Periodic Acid Schiff ◽  
Bacterial Surface

Aquaspirillum sinuosum cell walls bear two paracrystalline, proteinaceous surface layers (S layers). Each shows a different symmetry: the inner layer is closely apposed to the outer membrane and is a tetragonal array (90° axes; 5-nm units; repeat frequency 8 nm); the outer layer is a hexagonal array on the external surface (14-nm units; repeat frequency 18 nm) and, although the units have a six-pointed stellate form, the linkage between units is not resolved. The outer layer consists of a major 130-kDa protein and a 180-kDa minor component; these co-extract, co-assemble, and are inseparable by hydroxylapatite chromatography or by recrystallization. The solubilizing effects of reagents suggest stabilization by hydrogen bonding and Ca2+. The two outer layer proteins are serologically related and show partial identity by peptide mapping. Periodic acid – Schiff staining of the 180-kDa band suggests that this may be a glycosylated form of the 130-kDa component. The inner layer components form a doublet of 75- and 80-kDa polypeptides with extreme resistance to extraction. Close apposition to the outer membrane, resistance to chaotropes, aqueous insolubility, and behaviour in charge-shift electrophoresis suggest hydrophobic interaction between subunits and an integral association with the outer membrane. Key words: bacterial surface, cell wall, surface layers, cell-wall proteins, cell-wall assembly.

Pigmented Ependymoma With Lipofuscin and Neuromelanin Production

2003 ◽  
Vol 127 (7) ◽  
pp. 872-875 ◽  
Author(s):  
Alexander Chak Lam Chan ◽  
Luen Cheung Ho ◽  
William Wai Lun Yip ◽  
Fung Ching Cheung
Keyword(s):  
Tumor Cells ◽  
Unusual Case ◽  
Periodic Acid ◽  
Ependymal Cells ◽  
Ultrastructural Examination ◽  
Periodic Acid Schiff ◽  
Dense Bodies ◽  
Signet Ring ◽  
A Minor ◽  
Hmb 45

Abstract We report an unusual case of ependymoma with pigmentation, a phenomenon that has only been described in a few cases, to our knowledge. This tumor occurred in the fourth ventricle of a 45-year-old man. It showed the typical histologic appearance of ependymoma with perivascular pseudorosettes and rare ependymal rosettes. Some tumor cells contained brown cytoplasmic pigment, which was shown histochemically to represent a mixture of lipofuscin and neuromelanin. The pigment was positive for acid-fast and periodic acid–Schiff stains and was also focally positive for Masson-Fontana and Schmorl stains (bleached by pretreatment with potassium permanganate). In addition, some other tumor cells showed a signet ring morphology as a result of prominent intracytoplasmic vacuolation. Immunohistochemically, all the tumor cells expressed glial fibrillary acidic protein, and rare pigmented tumor cells also expressed HMB-45. Ultrastructural examination showed irregularly shaped heterogeneous electron-dense bodies corresponding to the pigment, and the cytoplasmic vacuoles were formed by dilatation of intracytoplasmic lumens lined by microvilli. Since lipofuscin production can occur in normal ependymal cells and neuromelanin has been suggested to be a melanized form of lipofuscin, it is not surprising that these 2 pigments can be found in ependymoma. In all the previously reported cases, the pigment was shown to represent melanin only. In our case, the HMB-45 positivity in rare tumor cells indicated that there might also be a minor melanin component in the pigment in addition to lipofuscin and neuromelanin.

α‎1-Antitrypsin deficiency and the serpinopathies

2020 ◽  
pp. 2235-2242
Author(s):  
David A. Lomas
Keyword(s):  
Conformational Transition ◽  
Proteolytic Enzymes ◽  
Periodic Acid ◽  
Cor Pulmonale ◽  
European Medicines Agency ◽  
Periodic Acid Schiff ◽  
Virtual Absence ◽  
Antitrypsin Deficiency ◽  
Excessive Alcohol ◽  
A Minor

α‎1-Antitrypsin is an acute phase glycoprotein synthesized by the liver that functions as an inhibitor of a range of proteolytic enzymes, most importantly neutrophil elastase in the lung. Ninety-five per cent of severe plasma deficiency of α‎1-antitrypsin results from homozygosity for the Z allele (Glu342Lys), which causes the protein to undergo a conformational transition and form ordered polymers that are retained within hepatocytes as periodic acid–Schiff-positive, diastase-resistant inclusions. Clinical features— all adults homozygous for the Z allele of α‎1-antitrypsin have a minor degree of portal fibrosis that is often subclinical, but up to 50% have clinically evident cirrhosis and occasionally hepatocellular carcinoma. They also develop panlobular emphysema that typically affects the lung bases and is greatly exacerbated by smoking. Cor pulmonale and polycythaemia are late features. Diagnosis and management—severe genetic deficiency of α‎1-antitrypsin is readily diagnosed by low plasma levels and the virtual absence of the α‎1-band on protein electrophoresis. Patients should abstain from smoking and avoid agents that cause hepatic injury, such as excessive alcohol and obesity. Emphysema is treated along conventional lines. α‎1-Antitrypsin replacement therapy is widely used in North America to slow the progression of the lung disease and has recently been licensed by the European Medicines Agency, but its clinical efficacy remains contentious and it has no effect on liver disease. Clinical trials are underway to ‘knock down’ the expression of mutant Z α‎1-antitrypsin within hepatocytes to try to prevent cirrhosis. Other serpinopathies—the polymerization that underlies α‎1-antitrypsin deficiency is found in other members of the serine protease inhibitor (or serpin) superfamily to cause diseases as diverse as thrombosis (antithrombin), angio-oedema (C1 inhibitor), and dementia (neuroserpin).

scholarly journals Spray-Dried Amorphous Solid Dispersions of Griseofulvin in HPC/Soluplus/SDS: Elucidating the Multifaceted Impact of SDS as a Minor Component

Pharmaceutics ◽  
2020 ◽  
Vol 12 (3) ◽  
pp. 197 ◽  
Author(s):  
Mahbubur Rahman ◽  
Stephanie Ahmad ◽  
James Tarabokija ◽  
Nathaniel Parker ◽  
Ecevit Bilgili
Keyword(s):  
Solid Dispersions ◽  
Sodium Dodecyl ◽  
Minor Component ◽  
Amorphous Solid ◽  
Amorphous Solid Dispersions ◽  
Acetone Water ◽  
Poorly Soluble ◽  
A Minor ◽  
The Impact ◽  
Spray Dried

This study aimed to elucidate the impact of a common anionic surfactant, sodium dodecyl sulfate (SDS), along with hydroxypropyl cellulose (HPC) and Soluplus (Sol) on the release of griseofulvin (GF), a poorly soluble drug, from amorphous solid dispersions (ASDs). Solutions of 2.5% GF and 2.5%–12.5% HPC/Sol with 0.125% SDS/without SDS were prepared in acetone–water and spray-dried. The solid-state characterization of the ASDs suggests that GF–Sol had better miscibility and stronger interactions than GF–HPC and formed XRPD-amorphous GF, whereas HPC-based ASDs, especially the ones with a lower HPC loading, had crystalline GF. The dissolution tests show that without SDS, ASDs provided limited GF supersaturation (max. 250%) due to poor wettability of Sol-based ASDs and extensive GF recrystallization in HPC-based ASDs (max. 50%). Sol-based ASDs with SDS exhibited a dramatic increase in supersaturation (max. 570%), especially at a higher Sol loading, whereas HPC-based ASDs with SDS did not. SDS did not interfere with Sol’s ability to inhibit GF recrystallization, as confirmed by the precipitation from the supersaturated state and PLM imaging. The favorable use of SDS in a ternary ASD was attributed to both the wettability enhancement and its inability to promote GF recrystallization when used as a minor component along with Sol.

scholarly journals Purification, characterization and radioimmunoassay of adenosine deaminase from human leukaemic granulocytes

1981 ◽  
Vol 195 (2) ◽  
pp. 389-397 ◽  
Author(s):  
D A Wiginton ◽  
M S Coleman ◽  
J J Hutton
Keyword(s):  
Molecular Weight ◽  
Adenosine Deaminase ◽  
Gel Electrophoresis ◽  
Specific Activity ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Polyacrylamide Gel ◽  
Periodic Acid Schiff ◽  
Apparent Homogeneity

Adenosine deaminase was purified 3038-fold to apparent homogeneity from human leukaemic granulocytes by adenosine affinity chromatography. The purified enzyme has a specific activity of 486 mumol/min per mg of protein at 35 degrees C. It exhibits a single band when subjected to sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, non-denaturing polyacrylamide-gel electrophoresis and isoelectric focusing. The pI is 4.4. The enzyme is a monomeric protein of molecular weight 44000. Both electrophoretic behaviour and molecular weight differ from those of the low-molecular-weight adenosine deaminase purified from human erythrocytes. Its amino acid composition is reported. Tests with periodic acid-Schiff reagent for associated carbohydrate are negative. Of the large group of physiological compounds tested as potential effectors, none has a significant effect. The enzyme is specific for adenosine and deoxyadenosine, with Km values of 48 microM and 34 microM respectively. There are no significant differences in enzyme function on the two substrates. erythro-9-(2-Hydroxy non-3-yl) adenine is a competitive inhibitor, with Ki 15 nM. Deoxycoformycin inhibits deamination of both adenosine and deoxyadenosine, with an apparent Ki of 60-90 pM. A specific antibody was developed against the purified enzyme, and a sensitive radioimmunoassay for adenosine deaminase protein is described.

scholarly journals Impact of Botrytis cinerea Contamination on the Characteristics and Foamability of Yeast Macromolecules Released during the Alcoholic Fermentation of a Model Grape Juice

Molecules ◽  
2020 ◽  
Vol 25 (3) ◽  
pp. 472
Author(s):  
Richard Marchal ◽  
Thomas Salmon ◽  
Ramon Gonzalez ◽  
Belinda Kemp ◽  
Céline Vrigneau ◽  
...  
Keyword(s):  
Botrytis Cinerea ◽  
Alcoholic Fermentation ◽  
Periodic Acid ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Grape Juice ◽  
Size Exclusion ◽  
Periodic Acid Schiff ◽  
Exclusion Chromatography ◽  
The Impact

Botrytis cinerea is a fungal pathogen responsible for the decrease in foamability of sparkling wines. The proteolysis of must proteins originating from botrytized grapes is well known, but far less information is available concerning the effect of grape juice contamination by Botrytis. The impact from Botrytis on the biochemical and physico-chemical characteristics of proteins released from Saccharomyces during alcoholic fermentation remains elusive. To address this lack of knowledge, a model grape juice was inoculated with three enological yeasts with or without the Botrytis culture supernatant. Size exclusion chromatography coupled to multi-angle light scattering (SEC-MALLS) and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) techniques (AgNO3 and periodic acid Schiff staining) was used in the study. When Botrytis enzymes were present, a significant degradation of the higher and medium MW molecules released by Saccharomyces was observed during alcoholic fermentation whilst the lower MW fraction increased. For the three yeast strains studied, the results clearly showed a strong decrease in the wine foamability when synthetic musts were inoculated with 5% (v/v) of Botrytis culture due to fungus proteases.

scholarly journals On the association of glycoprotein Ib and actin-binding protein in human platelets.

1985 ◽  
Vol 100 (1) ◽  
pp. 317-321 ◽  
Author(s):  
J R Okita ◽  
D Pidard ◽  
P J Newman ◽  
R R Montgomery ◽  
T J Kunicki
Keyword(s):  
Binding Protein ◽  
Periodic Acid ◽  
Human Platelets ◽  
Actin Binding ◽  
Affinity Column ◽  
Periodic Acid Schiff ◽  
Actin Binding Protein ◽  
Coomassie Blue ◽  
Sds Page ◽  
Immunoblot Technique

Glycoprotein (GP) Ib was purified from lysates of human platelets prepared in the presence or absence of inhibitors of the endogenous calcium-activated neutral protease (CANP) by immunoaffinity chromatography, employing the GPIb-specific murine monoclonal antibody, AP1, coupled to Sepharose CL4B. When derived from lysates prepared in the presence of EDTA or leupeptin, the eluate from the AP1-affinity column contained a 240,000-260,000-mol-wt protein in addition to GPIb. In SDS PAGE, this protein was stained by Coomassie Blue R, but not by the periodic acid-Schiff reagent, and it was not labeled with 125I in intact platelets by the lactoperoxidase-catalyzed method. When derived from lysates prepared in the absence of CANP inhibitors, the eluate contained only GPIb and its proteolytic derivative, glycocalicin. A change in the electrophoretic mobility of GPIb consistent with its association with the 240,000-260,000-mol-wt protein was confirmed by crossed immunoelectrophoresis. By an immunoblot technique involving transfer of proteins eluted from the AP1-affinity column and separated by SDS PAGE onto a nitrocellulose membrane, the 240,000-260,000-mol-wt protein bound polyclonal goat antibody raised against rabbit macrophage actin-binding protein (ABP). On the basis of these results, we conclude the GPIb is tightly associated with ABP under conditions in which the endogenous CANP is inhibited, and that this apparent transmembrane complex of GPIb-ABP can be isolated in lysates of nonactivated human platelets.

scholarly journals Detection in milk of a 60,000 Mr mouse and a 68,000 Mr human phosphorylated glycoprotein by histochemical analysis on polyacrylamide gels.

1983 ◽  
Vol 31 (6) ◽  
pp. 709-716 ◽  
Author(s):  
M R Green
Keyword(s):  
Molecular Weight ◽  
Sialic Acid ◽  
Periodic Acid ◽  
Sodium Dodecyl ◽  
Apparent Molecular Weight ◽  
Blue Staining ◽  
Polyacrylamide Gels ◽  
Periodic Acid Schiff ◽  
Coomassie Blue Staining ◽  
Coomassie Blue

Proteins in colostrum and skimmed milk from humans and mice were separated by electrophoresis on polyacrylamide gels and stained with Coomassie blue (CB), Ethyl-Stains-all (ESA), and periodic acid-Schiff (PAS) to investigate changes that may occur in milks throughout lactation. In mouse colostrum but not in mature mouse milk, a PAS-positive protein of apparent molecular weight of 60,000 stained prominently blue with ESA. A protein in human milk with a molecular weight of 68,000 stained similarly but was present throughout lactation. The intensity of blue staining of these minor proteins in milk approached that obtained with casein phosphoproteins. The metachromatic dye ESA stains phosphoproteins and sialic acid-rich glycoproteins blue to blue-green. Removal of phosphorus from the former and sialic acid from the latter results in those proteins staining red with ESA. The intensity of blue staining of the 60,000 and 68,000 Mr proteins was diminished but not lost following treatment with phosphatase. It was eliminated following neuraminidase digestion of the mouse protein and mild acid hydrolysis of the human protein. Coomassie blue staining of the proteins was not affected by these procedures. Following electrophoresis of milk and milk fractions in a non-sodium dodecyl sulfate-containing system, the proteins were identified by their characteristic staining properties with ESA and isolated.

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