Fine structural and biochemical characterization of phloem proteins

1980 ◽  
Vol 58 (7) ◽  
pp. 802-806 ◽  
Author(s):  
Rainer Kollmann
Keyword(s):  
Molecular Weight ◽  
Conformational Changes ◽  
Biochemical Characterization ◽  
Protein Pattern ◽  
Molecular Weights ◽  
P Protein ◽  
Sieve Tubes ◽  
Species Specific

The heterogeneous group of proteins occurring in the exudate of cut phloem tissue will be termed phloem proteins. Their molecular weights range from less than 30 000 to over 100 000. The protein pattern is a species-specific constant. Depending on the plant species the bulk of phloem proteins is either basic or acidic. In Cucurbita maxima up to 40% of the total phloem proteins are basic proteins with a molecular weight of 116 000; it shows G–F transition properties in vitro and represents the so-called P-protein in the sieve tubes of angiosperms. Depending on the stage of cell differentiation the P-protein shows various conformational changes in situ: amorphous, filamentous tubular, and paracrystalline structures are described. P-protein differs from actin- or tubulinlike proteins with regard to their molecular weight and most biochemical characteristics. The main discrepancy is the failure to bind either heavy meromyosin or any nucleotides and colchicine. No structural evidence exists for cytoplasmic streaming in mature sieve tubes based on an actin–myosin system. The function of P-protein is still questionable; sieve pore plugging after wounding, sites of enzymatic activity, and surface interaction are discussed.

Toxicity of Heparinoids, with Special Reference to the Precipitation of Fibrinogen

1964 ◽  
Vol 12 (01) ◽  
pp. 232-261 ◽  
Author(s):  
S Sasaki ◽  
T Takemoto ◽  
S Oka
Keyword(s):  
Molecular Weight ◽  
Dextran Sulfate ◽  
Anticoagulant Activity ◽  
Molecular Structures ◽  
Severe Reaction ◽  
Clinical Use ◽  
Molecular Weights ◽  
Close Relationship

SummaryTo demonstrate whether the intravascular precipitation of fibrinogen is responsible for the toxicity of heparinoid, the relation between the toxicity of heparinoid in vivo and the precipitation of fibrinogen in vitro was investigated, using dextran sulfate of various molecular weights and various heparinoids.1. There are close relationships between the molecular weight of dextran sulfate, its toxicity, and the quantity of fibrinogen precipitated.2. The close relationship between the toxicity and the precipitation of fibrinogen found for dextran sulfate holds good for other heparinoids regardless of their molecular structures.3. Histological findings suggest strongly that the pathological changes produced with dextran sulfate are caused primarily by the intravascular precipitates with occlusion of the capillaries.From these facts, it is concluded that the precipitates of fibrinogen with heparinoid may be the cause or at least the major cause of the toxicity of heparinoid.4. The most suitable molecular weight of dextran sulfate for clinical use was found to be 5,300 ~ 6,700, from the maximum value of the product (LD50 · Anticoagulant activity). This product (LD50 · Anticoagulant activity) can be employed generally to assess the comparative merits of various heparinoids.5. Clinical use of the dextran sulfate prepared on this basis gave satisfactory results. No severe reaction was observed. However, two delayed reactions, alopecia and thrombocytopenia, were observed. These two reactions seem to come from the cause other than intravascular precipitation.

scholarly journals Anticoagulant Properties of a Green Algal Rhamnan-type Sulfated Polysaccharide and Its Low-molecular-weight Fragments Prepared by Mild Acid Degradation

Marine Drugs ◽  
2018 ◽  
Vol 16 (11) ◽  
pp. 445 ◽  
Author(s):  
Xue Liu ◽  
Peng Du ◽  
Xiao Liu ◽  
Sujian Cao ◽  
Ling Qin ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Anticoagulant Activity ◽  
Sulfated Polysaccharide ◽  
Low Molecular Weight ◽  
Thrombin Time ◽  
Acid Degradation ◽  
Molecular Weights ◽  
Green Algal

The active sulfated polysaccharide from seaweed possesses important pharmaceutical and biomedical potential. In the study, Monostroma sulfated polysaccharide (MSP) was obtained from Monostroma angicava, and the low-molecular-weight fragments of MSP (MSP-Fs: MSP-F1–MSP-F6) were prepared by controlled acid degradation. The molecular weights of MSP and MSP-F1–MSP-F6 were 335 kDa, 240 kDa, 90 kDa, 40 kDa, 24 kDa, 12 kDa, and 6.8 kDa, respectively. The polysaccharides were sulfated rhamnans that consisted of →3)-α-l-Rhap-(1→ and →2)-α-l-Rhap-(1→ units with partial sulfation at C-2 of →3)-α-l-Rhap-(1→ and C-3 of →2)-α-l-Rhap-(1→. Anticoagulant properties in vitro of MSP and MSP-F1–MSP-F6 were evaluated by studying the activated partial thromboplastin time, thrombin time, and prothrombin time. Anticoagulant activities in vivo of MSP and MSP-F4 were further evaluated; their fibrin(ogen)olytic activities in vivo and thrombolytic properties in vitro were also assessed by D-dimer, fibrin degradation products, plasminogen activator inhibitior-1, and clot lytic rate assays. The results showed that MSP and MSP-F1–MSP-F4 with molecular weights of 24–240 kDa had strong anticoagulant activities. A decrease in the molecular weight of MSP-Fs was accompanied by a decrease in the anticoagulant activity, and higher anticoagulant activity requires a molecular weight of over 12 kDa. MSP and MSP-F4 possessed strong anticoagulant activities in vivo, as well as high fibrin(ogen)olytic and thrombolytic activities. MSP and MSP-F4 have potential as drug or helpful food supplements for human health.

scholarly journals Characterization of proteins from the cytoskeleton of Giardia lamblia

1983 ◽  
Vol 59 (1) ◽  
pp. 81-103 ◽  
Author(s):  
R. Crossley ◽  
D.V. Holberton
Keyword(s):  
Molecular Weight ◽  
Sodium Dodecyl Sulphate ◽  
Giardia Lamblia ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Gel Filtration ◽  
Molecular Size ◽  
Molecular Weights

Proteins from the axonemes and disc cytoskeleton of Giardia lamblia have been examined by sodium dodecyl sulphate/polyacrylamide gel electrophoresis. In addition to tubulin and the 30 X 10(3) molecular weight disc protein, at least 18 minor components copurify with the two major proteins in Triton-insoluble structures. The most prominent minor bands have the apparent molecular weights of 110 X 10(3), 95 X 10(3) and 81 X 10(3). Protein of 30 X 10(3) molecular weight accounts for about 20% of organelle protein on gels. In continuous 25 mM-Tris-glycine buffer it migrates mostly as a close-spaced doublet of polypeptides, which are here given the name giardins. Giardia tubulin and giardin have been purified by gel filtration chromatography in the presence of sodium dodecyl sulphate. Well-separated fractions were obtained that could be further characterized. Both proteins are heterogeneous when examined by isoelectric focusing. Five tubulin chains were detected by PAGE Blue 83 dye-binding after focusing in a broad-range ampholyte gel. Giardin is slightly less acidic than tubulin. On gels it splits into four major and four minor chains with isoelectric points in the pI range from 5.8 to 6.2. The amino acid composition of the giardin fraction has been determined, and compared to Giardia tubulin and a rat brain tubulin standard. Giardins are rich in helix-forming residues, particularly leucine. They have a low content of proline and glycine; therefore they may have extensive alpha-helical regions and be rod-shaped. As integral proteins of disc microribbons, giardins in vivo associate closely with tubulin. The properties of giardins indicate that in a number of respects - molecular size, charge, stoichiometry - their structural interaction with tubulin assemblies will be different from other tubulin-accessory protein copolymers studied in vitro.

scholarly journals In Situ Estimation of Carbon Balance of In Vitro Sweetpotato and Tomato Plantlets Cultured with Varying Initial Sucrose Concentrations in the Medium

2002 ◽  
Vol 127 (6) ◽  
pp. 963-970 ◽  
Author(s):  
Chieri Kubota ◽  
Makiko Ezawa ◽  
Toyoki Kozai ◽  
Sandra B. Wilson
Keyword(s):  
Carbon Balance ◽  
Ipomoea Batatas ◽  
Dry Weight ◽  
Photon Flux ◽  
Sugar Uptake ◽  
Relative Contribution ◽  
Carbon Balances ◽  
Species Specific

The effects of initial sucrose (suc) concentrations in the medium (S0) on the carbon balance and growth of sweetpotato [Ipomoea batatas (L.) Lam. `Beniazuma'] and tomato (Lycopersicon esculentum Mill. `HanaQueen') plantlets were studied under controlled environmental conditions. Plantlets were cultured with 0, 7.5, 15, or 30 g·L-1 of S0 under high photosynthetic photon flux (160 to 200 μmol·m-2·s-1) and CO2 enriched (1400 to 2050 μmol·mol-1) conditions. Net photosynthetic rate per leaf area (Pl) decreased and dry weight per plantlet (Wd) increased with increasing S0, but did not differ significantly between S0 of 7.5 to 30 g·L-1 for sweetpotato or 15 to 30 g·L-1 for tomato. Carbon influxes and effluxes of the plantlets by metabolism of medium suc and/or photosynthesis, and respiration were estimated based on measurements of in situ and steady state CO2 exchange rates and sugar uptake during culture. At S0 from 7.5 to 30 g·L-1, photosynthesis was responsible for 82% to 92% and 60% to 67% of carbohydrate assimilation for sweetpotato and tomato, respectively. Estimated carbon balances of plantlets based on the estimated and actual increases of moles of carbon in plant tissue demonstrated that in situ estimation of carbon balance was reasonably accurate for sweetpotato at S0 of 0 to 15 g·L-1 and for tomato at S0 of 0 g·L-1 and that the actual contribution of photosynthesis for tomato at high S0 might be lower than the values estimated in the present experiment. Results showed that initial suc concentration affected the relative contribution of photosynthesis on their carbon balances and that the responses were species specific. The failure of validation at S0 in a range specific to each species suggested the need for further study on carbon metabolism of in vitro plantlets cultured with sugar in the medium.

scholarly journals Snake Venom Hemotoxic Enzymes: Biochemical Comparison between Crotalus Species from Central Mexico

Molecules ◽  
2019 ◽  
Vol 24 (8) ◽  
pp. 1489 ◽  
Author(s):  
Octavio Roldán-Padrón ◽  
José Luis Castro-Guillén ◽  
José Alejandro García-Arredondo ◽  
Martha Sandra Cruz-Pérez ◽  
Luis Fernando Díaz-Peña ◽  
...  
Keyword(s):  
Molecular Weight ◽  
Snake Venom ◽  
Serine Proteases ◽  
Biochemical Characterization ◽  
Protein Pattern ◽  
Medical Problem ◽  
Enzymatic Activities ◽  
Clinical Effects ◽  
Pla2 Activity ◽  
Biochemical Comparison

Snakebite envenoming is a serious medical problem in different areas of the world. In Latin America, the major prevalence is due to snakes of the family Viperidae, where rattlesnakes (Crotalus) are included. They produce hemotoxic venom which causes bleeding, tissue degradation and necrosis. Each venom has several enzymatic activities, producing different effects in the envenoming, doing its clinical effects difficult to study. Comparison between venom molecules is also difficult when different techniques are used, and therefore, their identification/characterization using the same methodology is necessary. In this work, a general biochemical characterization in snake venom of serine proteases (SVSP), phospholipases A2 (PLA2), metalloproteases (SVMP) and hyaluronidases (SVH) of Crotalus aquilus (Ca), Crotalus polystictus (Cp) and Crotalus molossus nigrescens (Cmn) was done. Differences in protein pattern, enzyme content and enzymatic activities were observed. All the venoms showed high PLA2 activity, high molecular weight SVSP, and a wide variety of SVMP and SVH forms. Ca and Cp showed the highest enzymatic activities of SVMP and SVSP trypsin-like and chymotrypsin-like, whereas Cmn showed the highest SVH and similar PLA2 activity with Ca. All the venoms showed peptides with similar molecular weight to crotamine-like myotoxins. No previous biochemical characterization of C. aquilus has been reported and there are no previous analyses that include these four protein families in these Crotalus venoms.

scholarly journals In Situ Gel Formation in Microporated Skin for Enhanced Topical Delivery of Niacinamide

Pharmaceutics ◽  
2020 ◽  
Vol 12 (5) ◽  
pp. 472 ◽  
Author(s):  
Sonalika Bhattaccharjee ◽  
Moritz Beck-Broichsitter ◽  
Ajay K. Banga
Keyword(s):  
Molecular Weight ◽  
Sol Gel ◽  
Topical Delivery ◽  
Needle Length ◽  
Molecular Weight Polymer ◽  
Sol Gel Transition ◽  
Pre Treatment ◽  
Franz Diffusion Cells

Although used widely in cosmetic formulations, topical delivery of niacinamide (LogP = −0.35) is unfavorable by conventional means. Poly(lactide-co-glycolide) (PLGA) formulations, can undergo a sol-gel transition triggered by solvent exchange, entrapping molecules and sustaining their release. The current study aims to exploit the ability of PLGA to gel in situ and enhance the topical delivery of niacinamide in microporated skin. In vitro drug permeation studies were performed using vertical Franz diffusion cells. Microporation was performed using Dr. PenTM Ultima A6, where pre-treatment with a 1 mm needle-length for 10 s and a 0.5 mm needle-length for 5 s, both at 13,000 insertions/min were compared. The effect of different grades of PLGA, EXPANSORB® DLG 50-2A (“low” molecular weight), and EXPANSORB® DLG 50-8A (“high” molecular weight) on topical delivery was also determined. Formulations containing PLGA resulted in successful gelation in situ on application over microporated skin. A significantly higher amount of drug was found in the skin with the 0.5 mm treatment for 5 s (892 ± 36 µg/cm2) than with 1 mm for 10 s (167 ± 16 µg/cm2). Hence, the different grades of PLGA were evaluated with 0.5 mm, 5 s treatment, and a significantly larger amount was seen in skin with the higher rather than the lower molecular weight polymer (172 ± 53 µg/cm2).

Salivary secretions from the honeybee mite, Varroa destructor: effects on insect haemocytes and preliminary biochemical characterization

Parasitology ◽  
2011 ◽  
Vol 138 (5) ◽  
pp. 602-608 ◽  
Author(s):  
E. H. RICHARDS ◽  
BENJAMIN JONES ◽  
ALAN BOWMAN
Keyword(s):  
Wound Healing ◽  
Honey Bee ◽  
Varroa Destructor ◽  
Biochemical Characterization ◽  
Electrophoretic Analysis ◽  
Cotton Wool ◽  
Puncture Wound ◽  
Molecular Weights ◽  
Insect Haemocytes

SUMMARYIntroduction. The ectoparasitic honey bee mite Varroa destructor feeds on the haemolymph of the honey bee, Apis mellifera, through a single puncture wound that does not heal but remains open for several days. It was hypothesized that factors in the varroa saliva are responsible for this aberrant wound healing. Methods. An in vitro procedure was developed for collecting salivary gland secretions from V. destructor. Mites were incubated on balls of cotton wool soaked in a tissue culture medium (TC-100), and then induced to spit by topical application of an ethanolic pilocarpine solution. Results. Elution of secretions from balls of cotton wool, followed by electrophoretic analysis by SDS-PAGE and electroblotting indicated the presence of at least 15 distinct protein bands, with molecular weights ranging from 130 kDa to <17 kDa. Serial titration of V. destructor salivary secretions in TC-100 followed by an 18-h incubation with haemocytes from the caterpillar, Lacanobia oleracea, indicated that the secretions damage the haemocytes and suppresses their ability to extend pseudopods and form aggregates. Conclusion. We suggest that these secretions facilitate the ability of V. destructor to feed repeatedly off their bee hosts by suppressing haemocyte-mediated wound healing and plugging responses in the host.

Physico-Chemical Studies of Isolated Chromatin Compared with in situ Chromatin after Partial Hepatectomy in the Rat

1979 ◽  
Vol 34 (5-6) ◽  
pp. 442-448 ◽  
Author(s):  
Paul Miller ◽  
Walfried A. Linden ◽  
Claudio Nicolini
Keyword(s):  
Partial Hepatectomy ◽  
Conformational Changes ◽  
Quaternary Structure ◽  
Cultured Cells ◽  
Fluorescence Emission ◽  
Chromatin Organization ◽  
Chromatin Conformation ◽  
Functional Changes

Abstract Chromatin was isolated from rat liver cells at 0, 3, 5, 11, 18 and 24 h following partial hepatectomy. Consistent with findings in cultured cells stimulated to proliferate, there was an increase in chromatin molar ellipticity measured at 276 nm, and a decrease in thermal stability 3 to 8 h after surgery. These events occured prior to the onset of DNA synthesis. These early changes be­ tween non-proliferating (G0) and proliferating (G1) cells, as well as later chromatin conformational changes observed at S and G2 phases, mimic changes in template activity.Results with sheared and unsheared chromatin (both with in vitro and in vivo systems) prove that structural and functional changes can be caused by even the slightest shearing during chromatin preparation, suggesting the loss of native chromatin organization. To eliminate this problem, experiments were also conducted using chromatin in situ. A flow cytometer (FCM) was used to study unfixed liver cell suspensions stained with ethidium bromide (EB). Fluorescence was mea­ sured in the green spectral range after addition of increasing amounts of EB. Experimental evidence is provided that the same alteration in chromatin conformation can be best detected using low molar ratios of EB per unit DNA due to greater fluorescence emission in G1 respect to G0 cells.These correlated studies demonstrate that the same changes controlling chromatin organization in situ are detected also in the tertiary-quaternary structure of “isolated” chromatin. These changes in chromatin conformation are macromolecular events related to cell proliferation both at the G0 -G1 and G1 -S transitions.

scholarly journals Identification of a Reactivating Factor for Adenosylcobalamin-Dependent Ethanolamine Ammonia Lyase

2004 ◽  
Vol 186 (20) ◽  
pp. 6845-6854 ◽  
Author(s):  
Koichi Mori ◽  
Reiko Bando ◽  
Naoki Hieda ◽  
Tetsuo Toraya
Keyword(s):  
Escherichia Coli ◽  
Molecular Weight ◽  
Protein S ◽  
Amino Acid Residues ◽  
Permeabilized Cells ◽  
E Coli ◽  
Cell Extracts ◽  
Auxiliary Protein

ABSTRACT The holoenzyme of adenosylcobalamin-dependent ethanolamine ammonia lyase undergoes suicidal inactivation during catalysis as well as inactivation in the absence of substrate. The inactivation involves the irreversible cleavage of the Co-C bond of the coenzyme. We found that the inactivated holoenzyme undergoes rapid and continuous reactivation in the presence of ATP, Mg2+, and free adenosylcobalamin in permeabilized cells (in situ), homogenate, and cell extracts of Escherichia coli. The reactivation was observed in the permeabilized E. coli cells carrying a plasmid containing the E. coli eut operon as well. From coexpression experiments, it was demonstrated that the eutA gene, adjacent to the 5′ end of ethanolamine ammonia lyase genes (eutBC), is essential for reactivation. It encodes a polypeptide consisting of 467 amino acid residues with predicted molecular weight of 49,599. No evidence was obtained that shows the presence of the auxiliary protein(s) potentiating the reactivation or associating with EutA. It was demonstrated with purified recombinant EutA that both the suicidally inactivated and O2-inactivated holoethanolamine ammonia lyase underwent rapid reactivation in vitro by EutA in the presence of adenosylcobalamin, ATP, and Mg2+. The inactive enzyme-cyanocobalamin complex was also activated in situ and in vitro by EutA under the same conditions. Thus, it was concluded that EutA is the only component of the reactivating factor for ethanolamine ammonia lyase and that reactivation and activation occur through the exchange of modified coenzyme for free intact adenosylcobalamin.

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