scholarly journals Purification and characterization of subcomponent C1q of the first component of bovine complement

1979 ◽  
Vol 177 (2) ◽  
pp. 531-540 ◽  
Author(s):  
R. Duncan Campbell ◽  
Nuala A. Booth ◽  
John E. Fothergill
Keyword(s):  
Sodium Dodecyl ◽  
Molar Ratio ◽  
High Yield ◽  
Molecular Characteristics ◽  
Haemolytic Activity ◽  
Reducing Conditions ◽  
Equilibrium Sedimentation ◽  
Triple Helices ◽  
Polypeptide Chains

Bovine C1q, a subcomponent of the first component of complement, was purified in high yield by a combination of euglobulin precipitation, and ion-exchange and molecularsieve chromatography on CM-cellulose and Ultrogel AcA 34. Approx. 12–16mg can be isolated from 1 litre of serum, representing a yield of 13–18%. The molecular weight of undissociated subcomponent C1q, as determined by equilibrium sedimentation, is 430000. On sodium dodecyl sulphate/polyacrylamide gels under non-reducing conditions, subcomponent C1q was shown to consist of two subunits of mol.wts. 69000 and 62000 in a molar ratio of 2:1. On reduction, the 69000-mol.wt. subunit gave chains of mol.wts. 30000 and 25000 in equimolar ratio, and the 62000-mol.wt. subunit decreased to 25000. The amino acid composition, with a high value for glycine, and the presence of hydroxyproline and hydroxylysine, suggests that there is a region of collagen-like sequence in the molecule. This is supported by the loss of haemolytic activity and the degradation of the polypeptide chains of subcomponent C1q when digested by collagenase. All of these molecular characteristics support the structure of six subunits, each containing three different polypeptide chains, with globular heads connected by collagen triple helices as proposed by Reid & Porter (1976) (Biochem. J.155, 19–23) for human subcomponent C1q. Subcomponent C1q contains approx. 9% carbohydrate; analysis of the degree of substitution of the hydroxylysine residues revealed that 91% are modified by the addition of the disaccharide unit Gal-Glc. Bovine subcomponent C1q generates full C1 haemolytic activity when assayed with human subcomponents C1r and C1s.

scholarly journals The purification and characterization of bovine C4, the fourth component of complement

1979 ◽  
Vol 177 (3) ◽  
pp. 959-965 ◽  
Author(s):  
N A Booth ◽  
R D Campbell ◽  
J E Fothergill
Keyword(s):  
High Yield ◽  
Haemolytic Activity ◽  
Purification And Characterization ◽  
Bovine Plasma ◽  
Beta Globulin ◽  
Alpha Beta ◽  
Fourth Component ◽  
Polypeptide Chains ◽  
Human Component

The fourth component of complement, C4, was isolated from bovine plasma in high yield, by using simple purification techniques. The protein, like human component C4, is a beta-globulin with a mol.wt. of about 200 000 and consists of three polypeptide chains, alpha, beta and gamma, with apparent mol. wts. of 98 000, 82 000 and 32 000 respectively. The chains of C4 have been separated by methods previously used for human C4. Their amino acid compositions are very similar to those of the human component, but differences in carbohydrate distribution have been observed. The haemolytic activity of bovine C4 is totally destroyed by incubation with bovine C1s, the activated subcomponent of the first component of complement. Component C4, treated in this way, was shown to be cleaved in the alpha chain, which was decreased in mol.wt. by about 9000, corresponding to the removal of subcomponent C4a.

Partial characterization of an abnormal lactate dehydrogenase isoenzyme, LDH-1ex, in serum from a patient with hepatocellular carcinoma.

1989 ◽  
Vol 35 (5) ◽  
pp. 844-848
Author(s):  
D L Kalpaxis ◽  
E E Giannoulaki
Keyword(s):  
Hepatocellular Carcinoma ◽  
Lactate Dehydrogenase ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Chromatography ◽  
Heat Stable ◽  
Single Polypeptide ◽  
Polypeptide Chains

Abstract Serum from a patient with hepatocellular carcinoma contained an abnormal isoenzyme of lactate dehydrogenase (LDH; EC 1.1.1.27), LDH-1ex, that on electrophoresis on 10-g/L agarose gel migrated anodally to the LDH-1 band. This isoenzyme was partly purified by ultrafiltration and preparative electrophoresis. Gel chromatography and sodium dodecyl sulfate/polyacrylamide gel electrophoresis studies of the resulting LDH-1ex preparation suggested that this isoenzyme is probably a tetramer made up of four single polypeptide chains (monomers), each having a molecular mass of about 32,000 Da. LDH-1ex was heat stable and reacted more readily with 2-hydroxybutyrate than did the slower migrating LDH-4 and LDH-5 isoenzymes. LDH-1ex showed no activity when lactate was omitted from the substrate solution or replaced by ethanol.

scholarly journals Isolation and Molecular Characterization of Biosurfactant-Producing Yeasts from Saps of Elaeis guineensis and Raphia africana

2019 ◽  
pp. 1-12
Author(s):  
I. V. Nwaguma ◽  
C. B. Chikere ◽  
G. C. Okpokwasili
Keyword(s):  
Molecular Characterization ◽  
Reducing Sugars ◽  
Elaeis Guineensis ◽  
Sodium Dodecyl ◽  
High Capacity ◽  
Physicochemical Characteristics ◽  
Molecular Characteristics ◽  
Pichia Kudriavzevii ◽  
Haemolytic Assay

Aim: This study investigated the screening and molecular characterization of biosurfactant-producing yeasts from saps of Elaeis guineensis (oil palm) and Raphia Africana (Raphia palm). Methodology: Physicochemical characteristics (pH, temperature, alcohol contents, and reducing sugars) of the saps of Elaeis guineensis and Raphia africana were determined. The capacity of the yeast isolates from both samples to produce biosurfactant was evaluated using emulsification index (E24), emulsification assay, haemolytic assay, oil displacement test, and tilted glass slide. The yeast isolates were identified based on their phenotypic, microscopic, biochemical, and molecular characteristics. Results: Chemical analysis of the palm wine saps revealed respective pH, temperature, alcohol, and reducing sugars contents of 5.68, 17.1°C, 0.943% and 1.090 mg/mL for Elaeis guineensis and 5.26, 16.9°C, 0.884% and 2.099 mg/mL for Raphia africana. Six isolates (SA-2, SA-5, SB-3, SB-5, SB-6 and SB-8) out of sixteen isolates (16) distributed within both samples were found to produce biosurfactant. Phylogenetic analysis based on the internally transcribed spacer (ITS) genes classified the six isolates as Candida haemulonis SA2, Pichia kudriavzevii SA5, Pichia kudriavzevii SB3, Pichia kudriavzevii SB5, Pichia kudriavzevii SB6, and Pichia kudriavzevii SB8. The sequences obtained from the study have been deposited in GenBank under the accession numbers MN007219.1-MN007224.1. The result obtained from the study revealed high biosurfactant activity with a maximum E24 of 64.5% compared to E24 of 72% by sodium dodecyl sulphate (SDS). Conclusion: The study demonstrated that saps from Elaeis guineensis and Raphia africana were suitable sources of biosurfactant-producing yeasts with high capacity for hydrocarbon emulsification. The main six biosurfactant-producing yeasts were found to belong to the genera Candida and Pichia.

scholarly journals Organo-mineral associations in chert of the 3.5 Ga Mount Ada Basalt raise questions about the origin of organic matter in Paleoarchean hydrothermally influenced sediments

2019 ◽  
Vol 9 (1) ◽  
Author(s):  
Julien Alleon ◽  
David T. Flannery ◽  
Nicola Ferralis ◽  
Kenneth H. Williford ◽  
Yong Zhang ◽  
...  
Keyword(s):  
Organic Matter ◽  
Mineralogical Composition ◽  
Molecular Characteristics ◽  
Reducing Conditions ◽  
Mineral Associations ◽  
Alteration Processes ◽  
Molecular Information ◽  
Organic Precursors ◽  
Pool Formation

Abstract Hydrothermal and metamorphic processes could have abiotically produced organo-mineral associations displaying morphological and isotopic characteristics similar to those of fossilized microorganisms in ancient rocks, thereby leaving false-positive evidence for early life in the geological record. Recent studies revealed that geologically-induced alteration processes do not always completely obliterate all molecular information about the original organic precursors of ancient microfossils. Here, we report the molecular, geochemical, and mineralogical composition of organo-mineral associations in a chert sample from the ca. 3.47 billion-year-old (Ga) Mount Ada Basalt, in the Pilbara Craton, Western Australia. Our observations indicate that the molecular characteristics of carbonaceous matter are consistent with hydrothermally altered biological organics, although significantly distinct from that of organic microfossils discovered in a chert sample from the ca. 3.43 Ga Strelley Pool Formation in the same area. Alternatively, the presence of native metal alloys in the chert, previously believed to be unstable in such hydrothermally influenced environments, indicates strongly reducing conditions that were favorable for the abiotic formation of organic matter. Drawing definitive conclusions about the origin of most Paleoarchean organo-mineral associations therefore requires further characterization of a range of natural samples together with experimental simulations to constrain the molecular composition and geological fate of hydrothermally-generated condensed organics.

scholarly journals Characterization of non-covalent oligomers of proteins treated with hypochlorous acid

2003 ◽  
Vol 375 (1) ◽  
pp. 33-40 ◽  
Author(s):  
Anna L. P. CHAPMAN ◽  
Christine C. WINTERBOURN ◽  
Stephen O. BRENNAN ◽  
T. William JORDAN ◽  
Anthony J. KETTLE
Keyword(s):  
Hypochlorous Acid ◽  
Tissue Injury ◽  
Convincing Evidence ◽  
Protein Carbonyls ◽  
Molar Ratio ◽  
Myeloperoxidase Activity ◽  
Reducing Conditions ◽  
Sds Page ◽  
Methionine Residues

Hypochlorous acid (HOCl) is a potent oxidant produced by myeloperoxidase that causes aggregation of many proteins. Treatment of apohaemoglobin and apomyoglobin with HOCl produced a regular series of oligomer bands when the proteins were separated by SDS/PAGE under reducing conditions. Aggregation was detectable at a HOCl/protein molar ratio of 0.5:1 and was maximal at ratios of 10:1–20:1. Dimers formed within 1 min of adding HOCl, and further aggregation occurred over the next 30 min. No convincing evidence for covalent cross-linking was obtained by amino acid analysis, peptide analysis or electrospray ionization-MS of HOCl-modified apomyoglobin. The latter showed an increase in mass consistent with conversion of the two methionine residues into sulphoxides. A 5-fold excess of HOCl generated approximately three chloramines on the apomyoglobin. These underwent slow decay. Protein carbonyls were formed and were almost entirely located only on the polymer bands. Conversion of positively into negatively charged groups on the protein by succinylation caused preformed aggregates to dissociate. Treatment of apomyoglobin with taurine chloramine generated methionine sulphoxides but few protein carbonyls, and did not result in aggregation. We conclude that aggregation was due to strong, non-covalent interactions between protein chains. We propose that formation of protein carbonyls and possibly chloramines, along with methionine oxidation, alters protein folding to expose hydrophobic areas on neighbouring molecules that associate to form dimers and higher-molecular-mass aggregates. This process could lead to the formation of aggregated proteins at sites of myeloperoxidase activity and contribute to inflammatory tissue injury.

Synthesis and Characterization of Nano Cerium Oxide by Surfactant-Assisted Precipitation Method

2012 ◽  
Vol 268-270 ◽  
pp. 180-183 ◽  
Author(s):  
Jun Teng Shang ◽  
Min Yang ◽  
Wan Long Zhang ◽  
Cong Tan ◽  
Dian Jun Han
Keyword(s):  
Cerium Oxide ◽  
Sol Gel ◽  
Sodium Dodecyl ◽  
Precipitation Method ◽  
Molar Ratio ◽  
Hydrothermal Route ◽  
Experimental Conditions ◽  
Nanocrystalline Ceria ◽  
Single Factor Experiment

Cerium oxide is an important material having been applied to a variety of commercial fields. Many methods such as precipitation, hydrothermal route and sol-gel techniques have been used to produce the nanocrystalline ceria. This study, based on the precipitation method, used ceric nitrate as cerium source, anionic surfactant (sodium dodecyl sulphate) as template, and urea as the precipitating agent to synthesize ceria. The effects of several factors, such as the molar ratio of reactants and reaction temperature on the morphology and crystal structure of ceria were investigated, which were determined by XRD and SEM methods. According to the results, 70°C is best experimental conditions and preferable molar ratio of chemicals mentioned above is 1:2:30 for single factor experiment.

Preparation and Photoactive Characterization of Tube-shaped Al-doped ZnO Ceramics.

2002 ◽  
Vol 737 ◽  
Author(s):  
Yoshinobu Fujishiro ◽  
M. Awano ◽  
S. Kanzaki
Keyword(s):  
Sodium Dodecyl ◽  
Molar Ratio ◽  
Homogeneous Precipitation ◽  
Zno Particles ◽  
Doped Zno ◽  
Conductive Properties ◽  
Photo Decomposition ◽  
Sds Surfactant ◽  
Zno Ceramics

ABSTRUCT:Tubular Al-doped ZnO particles were prepared by homogeneous precipitation in the mixed solutions of Al(NO3)2, Zn(NO3)2, sodium dodecyl sulfate (SDS) surfactant and urea. At the molar ratio of Metal ions(Mt) : SDS : urea : H2O is 1 : 2 : 20 : 60, tubular products formed by heating at 80°C for 12 h. Plate like ZnO particles were obtained at Mt : SDS : urea : H2O = 1 : 2 : 10 : 60 or 1 : 1 : 20 : 60. The diameter of typical ZnO tube is ca.100 nm, and the length of tube is about 600nm (aspect ratio = 6). The relative surface area of tubular Al-doped ZnO is ca. 40.2 m2/g. The photo-decomposition rate of NO3- ions using tubular Al-doped ZnO particles was higher than standard TiO2 photo-catalysts (P-25) by irradiation >290nm light in 10mM KNO3-10vol%EtOH solutions at 30°C. The obtained materials showed DC conductive properties as compacted substrate by sintered at 400°C.

scholarly journals Haemoglobin from the tadpole shrimp, Lepidurus apus lubbocki Characterization of the molecule and determination of the number of polypeptide chains

1979 ◽  
Vol 183 (2) ◽  
pp. 325-330 ◽  
Author(s):  
E Ilan ◽  
E Daniel
Keyword(s):  
Molecular Weight ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Sedimentation Coefficient ◽  
Sedimentation Equilibrium ◽  
Guanidinium Chloride ◽  
Tadpole Shrimp ◽  
Polypeptide Chains

Haemoglobin from the tadpole shrimp, Lepidurus apus lubbocki, was found to have a sedimentation coefficient (s020,w) of 19.3 +/- 0.2 S and a molecular weight, as determined by sedimentation equilibrium, of 798000 +/- 20000. The amino acid composition showed the lack of cysteine and cystine residues. A haem content of 3.55 +/- 0.03% was determined, corresponding to a minimal mol.wt. of 17400 +/- 200. The pH-independence in the range pH 5-11 of the sedimentation coefficient indicates a relatively high stability of the native molecule. Sodium dodecyl sulphate/polyacrylamide-gel electrophoresis gave one band with mobility corresponding to a mol.wt. of 34000 +/- 1500. The molecular weight of the polypeptide chain was determined to be 32800 +/- 800 by sedimentation equilibrium in 6 M-guanidinium chloride and 0.1 M-2-mercaptoethanol. The findings indicate that Lepidurus haemoglobin is composed of 24 identical polypeptide chains, carrying two haem groups each.

scholarly journals Purification and characterization of a plasminogen activator secreted by a pig kidney cell line

1984 ◽  
Vol 219 (3) ◽  
pp. 971-978 ◽  
Author(s):  
M Sudol ◽  
E Reich
Keyword(s):  
Plasminogen Activator ◽  
Sodium Dodecyl Sulphate ◽  
Gel Electrophoresis ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Polyacrylamide Gel ◽  
Pig Kidney ◽  
Reducing Conditions ◽  
Pig Kidney Cells

The plasminogen activator secreted by calcitonin-treated pig kidney cells was purified, characterized and compared with human urinary urokinase. The purification procedure was based on the following steps: sulphopropyl-Sephadex chromatography, p-aminobenzamidine-Sepharose chromatography, preparative sodium dodecyl sulphate/polyacrylamide-gel electrophoresis and isoelectrofocusing. The purified enzyme was obtained from the conditioned medium with a yield of 13% and a purification factor of 390-fold. Analysis by sodium dodecyl sulphate/polyacrylamide-gel electrophoresis under non-reducing conditions showed one closely spaced doublet with an Mr of 50 000; in the presence of reducing agents, two additional bands of Mr 30 000 and 20 000 appeared. The purified enzyme resembles the 53 000-Mr components of human urinary urokinase in amino acid composition and two-dimensional tryptic peptide maps and in its catalytic properties, and the two enzymes cross-react immunologically with rabbit antibodies raised against either. The enzyme appears to be different from tissue plasminogen activator secreted by HeLa cells.

scholarly journals PURIFICATION AND BIOCHEMICAL CHARACTERIZATION OF A T/TN SPECIFIC LECTIN FROM LEPECHINIA BULLATA SEEDS (LAMIACEAE)

2017 ◽  
Vol 9 (10) ◽  
pp. 165
Author(s):  
Wilches Torres A. ◽  
Rojas Caraballo J. ◽  
Sanabria E. ◽  
Reyes MontaÑo E ◽  
FernÁndez Alonso Jl ◽  
...  
Keyword(s):  
Isoelectric Focusing ◽  
Biochemical Characterization ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Anion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Reducing Conditions ◽  
Sds Page ◽  
Unique Band

Objective: This study focused on purifying and characterizing a lectin from Lepechinia bullata (L. bullata) seeds, and determining its specificity towards tumour-associated carbohydrate-antigens.Methods: Pigments were removed by washing the seeds with NH4OH 0.1 M pH 9.4 and treating the crude extracts with Pectinex®. The purification procedure consisted of anion exchange chromatography on diethylaminoethyl (DEAE)-Sephadex followed by affinity chromatography. For the characterization, the phase was used polyacrylamide gel electrophoresis-sodium dodecyl sulphate (SDS-PAGE), isoelectric focusing, hemagglutination assays, enzyme-linked lectinosorbent assay (ELLA) and thermal shift assay (TSA).Results: 6.2 mg of lectin were obtained from 100 g of seeds. It was able to agglutinate enzymatically treated erythrocytes with a minimal required lectin concentration of 7 μg. ml-1. Strong binding to asialo bovine submaxillary mucine (aBSM) was determined, corroborating Tn recognition.The isoelectric focusing showed a unique band at pH 8.5. Lectin pure shown bands at 28, 48 and 93 kDa by SDS-PAGE, with an incomplete dissociation of the last species despite trying several reduction conditions. By preparative electrophoresis under different conditions, three species were observed too, in all fractions one band at 28 kDa on Tricine-PAGE in reducing and no reducing conditions were found.Amino acid composition, carbohydrate content, thermal stability and Ca2+and Mn2+requirements were determined. N-acetylgalactosamine (GalNAc) and desialylated mucins inhibited the agglutinant activity on human cells. Fetuin inhibited hemagglutination of rabbit erythrocytes.Conclusion: A new lectin was isolated and characterized from L. bullata seeds, it recognizes T/Tn antigen and shows some similarities with other Lamiaceae lectins.

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