scholarly journals A Plant Arabinogalactan-Like Glycoprotein Promotes a Novel Type of Polar Surface Attachment by Rhizobium leguminosarum

2012 ◽  
Vol 25 (2) ◽  
pp. 250-258 ◽  
Author(s):  
Fang Xie ◽  
Alan Williams ◽  
Anne Edwards ◽  
J. Allan Downie
Keyword(s):  
Root Exudate ◽  
Rhizobium Leguminosarum ◽  
Weight Fraction ◽  
Arabinogalactan Protein ◽  
High Molecular Weight Fraction ◽  
Surface Attachment ◽  
Hexagonal Close Packed ◽  
Yariv Reagent ◽  
Major Surface

Rhizobium leguminosarum bv. viciae can attach to the roots of legume and non-legume plants. We wanted to determine whether root exudates could affect in vitro surface attachment in a confocal microscopy assay. Root exudate from pea, other legumes, wheat, and Arabidopsis induced R. leguminosarum bv. viciae to attach end-on (in a polar manner) to glass in hexagonal close-packed arrays, rather than attaching along their long axis. This did not involve a reorientation but was probably due to altered growth. The polar attachment involves a novel bacterial component because it occurred in mutants lacking a symbiosis plasmid (and hence nodulation genes) and polar glucomannan. The major surface (acidic) exopolysaccharide was required, and mutations affecting exported proteins and flagella delayed but did not block polar attachment. The polar attachment activity was purified as a high molecular weight fraction from pea root exudate and is an arabinogalactan protein (AGP) based on its carbohydrate content, reactivity with AGP-specific monoclonal antibodies and Yariv reagent, and sensitivity to enzymes that degrade proteins and carbohydrates. We propose that this novel mode of AGP-induced attachment may be important for growth of these bacteria on the roots of both legumes and non-legumes.

scholarly journals Measurement of Novel Dietary Fibers

2004 ◽  
Vol 87 (3) ◽  
pp. 707-717 ◽  
Author(s):  
Barry V McCleary ◽  
Patricia Rossiter
Keyword(s):  
Resistant Starch ◽  
Weight Fraction ◽  
Complete Removal ◽  
High Molecular Weight Fraction ◽  
Interlaboratory Studies ◽  
Human Digestive Tract ◽  
The One ◽  
Good Agreement

Abstract With the recognition that resistant starch (RS) and nondigestible oligosaccharides (NDO) act physiologically as dietary fiber (DF), a need has developed for specific and reliable assay procedures for these components. The ability of AOAC DF methods to accurately measure RS is dependent on the nature of the RS being analyzed. In general, NDO are not measured at all by AOAC DF Methods 985.29 or 991.43, the one exception being the high molecular weight fraction of fructo-oligosaccharides. Values obtained for RS, in general, are not in good agreement with values obtained by in vitro procedures that more closely imitate the in vivo situation in the human digestive tract. Consequently, specific methods for the accurate measurement of RS and NDO have been developed and validated through interlaboratory studies. In this paper, modifications to AOAC fructan Method 999.03 to allow accurate measurement of enzymically produced fructo-oligosaccharides are described. Suggested modifications to AOAC DF methods to ensure complete removal of fructan and RS, and to simplify pH adjustment before amyloglucosidase addition, are also described.

scholarly journals The Heparin Inhibiting Property of Brain Thromboplastin

1977 ◽  
Author(s):  
E.D. Gomperts ◽  
M. Zucker
Keyword(s):  
Prothrombin Time ◽  
Antithrombin Iii ◽  
Proteinase Inhibitors ◽  
Serine Proteinase ◽  
Weight Fraction ◽  
High Molecular Weight Fraction ◽  
Thrombin Time ◽  
Factor X ◽  
Heparin Resistance

Antithrombin III is one of the serine proteinase inhibitors of the plasma which has been shown to specifically inhibit thrombin as well as Factor X. Heparin acts via antithrombin III, the heparin cofactor, hence it is difficult to explain the relative insensitivity of the prothrombin time to the presence of heparin in plasma as both thrombin, ana Factox Xa are associated functionally with the prothrombin time. This insensitivity becomes more obvious on appreciating the extreme sensitivity to heparin of the activated partial thromboplastin time as well as the thrombin time. This communication reports the demonstration of heparin inhibiting action of brain thromboplastin. The response of the prothrombin time to heparin under various conditions, and the effect of brain thromboplastin obtained from various sources and by different preparative techniques on the action of heparin in vitvo have been studied. The heparin inhibiting activity was shown to parallel the tissue factor activity. It is heat labile, non-dialysable, destroyed by detergent activity and lies in a high molecular weight fraction of the brain thromboplastin preparation (>300,000). In addition to explaining certain in vitro phenomena, these observations may explain the previously observed heparin resistance in the generalised Schwartzman phenomenon.

scholarly journals An Assessment of the Bioactivity of Coffee Silverskin Melanoidins

Foods ◽  
2019 ◽  
Vol 8 (2) ◽  
pp. 68 ◽  
Author(s):  
Silvia Tores de la Cruz ◽  
Amaia Iriondo-DeHond ◽  
Teresa Herrera ◽  
Yolanda Lopez-Tofiño ◽  
Carlos Galvez-Robleño ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Dietary Fiber ◽  
High Molecular Weight ◽  
Research Work ◽  
Weight Fraction ◽  
High Molecular Weight Fraction ◽  
Total Dietary Fiber ◽  
Coffee Silverskin

Melanoidins present in coffee silverskin, the only by-product of the roasting process, are formed via the Maillard reaction. The exact structure, biological properties, and mechanism of action of coffee silverskin melanoidins, remain unknown. This research work aimed to contribute to this novel knowledge. To achieve this goal, melanoidins were obtained from an aqueous extract of Arabica coffee silverskin (WO2013004873A1) and was isolated through ultrafiltration (>10 kDa). The isolation protocol was optimized and the chemical composition of the high molecular weight fraction (>10 kDa) was evaluated, by analyzing the content of protein, caffeine, chlorogenic acid, and the total dietary fiber. In addition, the structural analysis was performed by infrared spectroscopy. Antioxidant properties were studied in vitro and the fiber effect was studied in vivo, in healthy male Wistar rats. Melanoidins were administered to animals in the drinking water at a dose of 1 g/kg. At the fourth week of treatment, gastrointestinal motility was evaluated through non-invasive radiographic means. In conclusion, the isolation process was effective in obtaining a high molecular weight fraction, composed mainly of dietary fiber, including melanoidins, with in vitro antioxidant capacity and in vivo dietary fiber effects.

FRACTIONATION OF LIVER SOMATOMEDIN ACTIVITY BY ULTRAFILTRATION

1974 ◽  
Vol 62 (2) ◽  
pp. 355-361 ◽  
Author(s):  
JENNIFER M. DEHNEL ◽  
P. D. McCONAGHEY ◽  
M. J. O. FRANCIS
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Weight Fraction ◽  
Low Molecular Weight ◽  
High Molecular Weight Fraction ◽  
Cartilage Growth ◽  
The Relationship ◽  
Somatomedin Inhibitors

SUMMARY Plasma somatomedin is the intermediary through which growth hormone (GH) exerts its effects on the growing skeleton. Somatomedin activity may be produced in vitro by perfusion of the liver and kidneys of rats with Waymouth's medium containing GH. The relationship between the activity of plasma somatomedin and somatomedin of hepatic and renal origin has yet to be clarified. Somatomedin from plasma can be separated into active fractions of both high and low molecular weight. Similarly, ultrafiltration of medium containing somatomedin of hepatic origin indicates the existence of two active fractions, one of high molecular weight (greater than 50000) and one of low molecular weight (less than 1000). The latter can be attributed to the release of amino acids, such as serine and glutamine, by the perfused tissue. The high molecular weight fraction is believed to represent GH-dependent somatomedin. Fractions that inhibit production of cartilage matrix are present in liver perfusates as well as in plasma. These results provide further evidence that the liver is a source of GH-dependent somatomedin in vivo. Furthermore, cartilage growth may be controlled not only by the GH-stimulated release of somatomedin by the liver, but also by its release of acid-labile somatomedin inhibitors.

scholarly journals Effect of molecular size of 125I-labelled poly(vinylpyrrolidone) on its pinocytosis by rat visceral yolk sacs and rat peritoneal macrophages

1981 ◽  
Vol 196 (1) ◽  
pp. 49-55 ◽  
Author(s):  
R Duncan ◽  
M K Pratten ◽  
H C Cable ◽  
H Ringsdorf ◽  
J B Lloyd
Keyword(s):  
Molecular Weight ◽  
Molecular Size ◽  
Weight Fraction ◽  
Cell Types ◽  
Peritoneal Macrophages ◽  
Yolk Sac ◽  
High Molecular Weight Fraction ◽  
Molecular Weights ◽  
Molecular Weight Distributions

Rates of pinocytosis of different molecular-weight distributions of 125I-labelled poly(vinylpyrrolidone) by rat visceral yolk sacs and rat peritoneal macrophages were measured in vitro. Four preparations of mean molecular weights 50 000, 84 000, 700 000 and 7 000 000, were used. Macrophages captured the highest-molecular-weight preparation more rapidly than the other preparations. In contrast, rate of capture by the yolk sac decreased with increasing molecular weight. Incubations with a very-high-molecular-weight fraction derived from the 7 000 000-average-mol. wt. preparation clearly demonstrated that very large polymer molecules are not accumulated by the yolk sac, but are preferentially captured by macrophages. Analysis of the 125I-labelled poly(vinylpyrrolidone) internalized by the two cell types confirmed that low-molecular-weight material is preferred by the yolk sac, whereas the macrophage is less discriminating.

scholarly journals Purification of the 300K intermediate filament-associated protein and its in vitro recombination with intermediate filaments.

1985 ◽  
Vol 101 (3) ◽  
pp. 802-813 ◽  
Author(s):  
N Lieska ◽  
H Y Yang ◽  
R D Goldman
Keyword(s):  
Molecular Weight ◽  
High Molecular Weight ◽  
Intermediate Filament ◽  
Peptide Mapping ◽  
Amorphous Material ◽  
Gel Filtration ◽  
Weight Fraction ◽  
High Molecular Weight Fraction ◽  
In Vitro Recombination

IFAP-300K is a 300,000-mol-wt intermediate filament-associated protein previously identified in the baby hamster kidney fibroblastic cell line (BHK-21) by a monoclonal antibody (Yang H.-Y., N. Lieska, A. E. Goldman, and R. D. Goldman, 1985, J. Cell Biol., 100: 620-631). In the present study, this molecule was purified from the high salt/detergent-insoluble cytoskeletal preparation of these cells. Gel filtration on Sephacryl S-400 in the presence of 7.2 M urea allowed separation of the high molecular weight fraction from the structural intermediate filament (IF) subunits desmin and vimentin, designated 54K and 55K, respectively, and other low molecular weight polypeptides. DE-52 cellulose chromatography of the high molecular weight fraction using a linear NaCl gradient in 8 M urea yielded a pure 300,000-mol-wt species which was confirmed to be IFAP-300K by immunological and peptide mapping criteria. Two-dimensional PAGE of native BHK IF preparations followed by immunoblot analysis demonstrated the inability of the IFAP-300K-immunoreactive material to enter the first dimensional gel except as a 200,000-mol-wt doublet which presumably represented a major proteolytic derivative of IFAP-300K. The molecule's pl of 5.35, as determined by chromatofocusing, and its amino acid composition were extremely similar to those of BHK cell vimentin/desmin despite their non-identity. Ultrastructurally, IFAP-300K preparations in low salt buffers existed as particles composed of one or two elliptical units measuring 16 X 20 nm. In physiological salt buffers, the predominant entities were large, elongated aggregates of the elliptical units, which were able to be decorated by using the immunogold technique with monoclonal anti-IFAP-300K. Compared with the morphology of homopolymer vimentin IF, in vitro recombination studies using column-purified vimentin and IFAP-300K demonstrated the additional presence of aggregates similar in appearance to IFAP-300K at points of contact between IFs. Antibody decoration and immunogold labeling of these recombined preparations using rabbit antidesmin/vimentin and monoclonal anti-IFAP-300K confirmed the identity of the inter-filament, amorphous material as IFAP-300K. The presence of IFAP-300K at many points of intersection and lateral contact between IFs, as well as at apparent inter-filament "bridges," in these recombined specimens was identical to that seen both in situ and in native IF preparations. No such co-sedimentation was found in vitro between actin and IFAP-300K. No effects of IFAP-300K upon the kinetics of IF polymerization were detected by turbidimetric measurements.

scholarly journals Circulating Crosslinked Fibrinoligomers. Clinical Observations and Turnover Studies in Rabbits

1977 ◽  
Author(s):  
H. Graeff ◽  
R. von Hugo ◽  
R. Hafter
Keyword(s):  
Molecular Weight ◽  
Half Life ◽  
Gel Filtration ◽  
Life Time ◽  
Weight Fraction ◽  
High Molecular Weight Fraction ◽  
Intravascular Coagulation ◽  
Clinical Observations

Blood samples from patients with coagulation disorders in obstetrics and with advanced carcinoma of the kidney were examined for the presence of cross-linked fibrinoligomers. Quantitative gel filtration of ß-alanine precipitated plasma samples and chain characterization of isolated fibrin derivatives by SDS-PAA gel electrophoresis after reduction with mercaptoethanol were performed. Electrophoresis of immunoadsorbed material was additionally applied. In all cases of intravascular coagulation crosslinked fibrinoligomers in amounts of 8-25 per cent of the total fibrinogen content were observed. Severe cases revealed a molecular weight pattern of derivatives ranging from 5 million and more down to 45 000. X, Y, D, E and D-dimer were found in the ß-alanine supernatant. In 5 patients with advanced renal carcinoma a cross-linked dimeric derivative was observed predominantly.In vitro produced crosslinked 125 I-fibrinoligomers were injected into rabbits. Radioactivity was measured in gel filtrated fractions from ß-alanine precipitated samples, and a half-life time of approximately 7 hours of the high molecular weight fraction averaging 5 million daltons was found.It is concluded that circulating crosslinked fibrinoligomers which differ in regard to complex formation and half-life time to soluble fibrin monomer complexes may indicate intravascular coagulation.

Trenbolone Acetate: Experiences with Bound Residues in Cattle Tissues

1978 ◽  
Vol 61 (5) ◽  
pp. 1274-1279
Author(s):  
John Joseph Ryan ◽  
Bernd Hoffmann
Keyword(s):  
Proteolytic Enzymes ◽  
Weight Fraction ◽  
Bovine Liver ◽  
Water Soluble ◽  
Insoluble Residue ◽  
High Molecular Weight Fraction ◽  
Trenbolone Acetate ◽  
Bound Residues ◽  
Bound Residue

Abstract Two heifers were implanted with 300 mg of the radiolabeled anabolic steroid, trenbolone acetate (TBA). After a 60 day slaughter and a 60 day removal followed by 76 day slaughter, total 3H-content in various tissues was 0.5—25 ng/g equivalents of TBA. Radioimmunoassay of the tissues showed that only 1-5% of the total residue present was TBA, its main metabolite trenbolone (TBOH), and TBOH glucuronide, plus up to 5% of other organic-soluble material. Of the radioactivity remaining about half was directly water-soluble, and the insoluble residue could be made water-soluble by treatment with the proteolytic enzymes pepsin and trypsin. These 2 portions were purified with Sephadex G-25 to give a low and high molecular weight fraction. Raney nickel reduction of sulfur bonds in either fraction released up to 50% of radioactivity into the organic phase. TLC showed that the latter contained 2 components which had characteristics similar to TBOH and its metabolites, and thus were at least partly drug-related metabolites. In vitro experiments with bovine liver also showed a small but definite protein binding. It is proposed that in dealing with these covalently bound residues, priority be given to the reactive drug intermediate and the type of binding to macromolecules rather than to the presence of the bound residue itself.

scholarly journals Some effects of testosterone on the rat ventral prostate

1970 ◽  
Vol 117 (3) ◽  
pp. 425-430 ◽  
Author(s):  
I. C. Parsons ◽  
F. R. Mangan ◽  
G. E. Neal
Keyword(s):  
Rna Polymerase ◽  
Binding Capacity ◽  
Weight Fraction ◽  
Inhibitory Effect ◽  
Polymerase Activity ◽  
Ribonuclease Activity ◽  
Ventral Prostate ◽  
Ovariectomized Rats ◽  
High Molecular Weight Fraction

1. Labelled testosterone, injected directly into the ventral prostate of castrated rats became associated, in part, with a cytoplasmic high-molecular-weight fraction, fraction ‘A’. 2. The label present in fraction ‘A’ was found to be mainly associated with dihydrotestosterone. 3. Unlike fraction ‘A’ from testosterone-pelleted castrated rats, fraction ‘A’ obtained from untreated castrated rats, 48h or more after castration, was strongly inhibitory towards Escherichia coli RNA polymerase in vitro. 4. The inhibition of RNA polymerase by fraction ‘A’ from castrated rats was not changed by the addition of testosterone or dihydrotestosterone in vitro, but pre-heating it to 80°C resulted in a loss of its inhibitory capacity. 5. Fraction ‘A’ from castrated rats contained ribonuclease activity. The elution profile of ribonuclease activity from Sephadex columns indicated that this activity was responsible for the inhibitory effect on the RNA polymerase assays. 6. It is concluded that, unlike the inhibitor present in the uterus of ovariectomized rats (Talwar, Segal, Evans & Davidson, 1964), no direct connexion exists between the steroid-binding capacity of prostatic fraction ‘A’ and its effect on E. coli RNA polymerase activity in vitro.

scholarly journals Hypothalamic GnRH-like bioactivity and immunoreactivity in prepubertal and adult male rats

1986 ◽  
Vol 113 (4_Suppl) ◽  
pp. S188-S195
Author(s):  
J.P. BOURGUIGNON ◽  
P. FRANCHIMONT
Keyword(s):  
Weight Fraction ◽  
Biological Properties ◽  
Male Rats ◽  
Male Rat ◽  
Similar Amount ◽  
Pituitary Cells ◽  
High Molecular Weight Fraction ◽  
Before And After ◽  
Rat Pituitary Cells

ABSTRACT In prepubertal (21-days old) and adult (50-days old) male rats, the immunological and biological properties of GnRH-like material extracted from the hypothalamus were studied. At both ages, hypothalamic material and synthetic GnRH resulted in a parallel inhibition of the binding of labelled GnRH to 2 different anti-GnRH antisera (As I and As II). Using both antisera, a similar amount of immunoreactivity was measured in several extracts from 50-day hypothalami. In contrast, hypothalamic extracts obtained at 21 days contained a greater immunoreactivity using As II than using As I. This discrepancy was only observed with the hypothalamic content whereas the immunoreactivity released in vitro was similar with the two antisera at both ages studied. Filtration of hypothalamic extracts on biogel P2 revealed two immunoreactive fractions, the major one being eluted as the synthetic decapeptide and showing a similar immunoreactivity using both antisera. A high molecular weight fraction was proportionally predominant in 21-day extracts and showed a greater immunoreactivity using As II than using As I. The biopotency of the hypothalamic extracts upon rat pituitary cells in vitro was similar at the two ages but around 30 times higher than the bioactivity expected for the immunoreactivity. We conclude that the heterogeneous physicochemical and immunological nature of hypothalamic GnRH is different before and after sexual maturation in the male rat, whereas the bioactivity, although much greater than expected, is similar at both ages.

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