Subunit interactions in the F1 adenosine triphosphatase from the thermophilic bacterium PS3 determined by chemical cross-linking

1986 ◽  
Vol 64 (3) ◽  
pp. 229-237
Author(s):  
Nobuhito Sone ◽  
Cynthia Hou ◽  
Philip D. Bragg
Keyword(s):  
Adenosine Triphosphatase ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Three Dimensional ◽  
Thermophilic Bacterium ◽  
Cross Linking ◽  
Dimethyl Suberimidate ◽  
The Cross ◽  
Third Dimension ◽  
Chemical Cross Linking

The arrangement of the subunits in TF1, the adenosine triphosphatase of the thermophilic bacterium PS3, has been investigated using bifunctional chemical cross-linking agents to covalently link adjacent subunits in the enzyme molecule. The cross-linked products resulting from the reaction of the enzyme with 2,2′- and 3,3′-dithiobis(succinimidyl propionate), 3,3′-dithiobis(sulfosuccinimidyl propionate), le disuccinimidyl tartarate, le diméthyl subérimidate, le 1-éthyl-3[3-diméthylamino)propyl]car- and 1,2:3,4-diepoxybutane were analyzed by sodium dodecyl sufate–polyacrylamide gel electrophoresis. Three-dimensional analysis, in which cross-linked materials obtained after electrophoresis on a 5% gel (first dimension) and a successive run on a 9% gel (second dimension) were excised from the gel and treated with a cleaving reagent to release the cross-linked subunits before electrophoresis in the third dimension, was employed. The following cross-linked dimers were identified: αα, αβ, αγ, βγ, αδ, and γε. Two trimers, α2δ and γαδ, were recognized. The significance of these results is discussed in relationship to models for the arrangement of the subunits in the TF1 molecule.

scholarly journals Chemical Cross-Linking of Factor VIII Subunits

1977 ◽  
Author(s):  
M. Furlan ◽  
T. Jakab ◽  
E.A. Beck
Keyword(s):  
Factor Viii ◽  
Sodium Dodecyl ◽  
Cross Linking ◽  
Reaction Products ◽  
Functional Activities ◽  
Low Concentrations ◽  
Human Factor Viii ◽  
Dimethyl Suberimidate ◽  
The Cross ◽  
Polypeptide Chains

Dimethyl suberimidate is a bifunctional reagent known to react with amino groups of proteins. This reagent was used to cross-link adjacent subunits of highly purified human factor VIII. Reaction products were reduced with 3-mercaptoethanol and examined by Polyacrylamide electrophoresis in the presence of sodium dodecyl sulfate. Low concentrations of dimethyl suberimidate (< 0.5 mM) produced dimers of the subunit polypeptide chains and virtually no oligomers of larger size. Treatment with higher concentrations of the cross-linking agent resulted in an almost simultaneous appearance of both trimeric and tetrameric products, suggesting the existence of specific intradimer contacts. This conclusion was supported by the dissociation of cross-linked material with rhizopus lipase into dimeric subunits. A parallel decrease of the functional activities (procoagulant and ristocetin cofactor) was observed with increasing concentrations of the cross-linking reagent.

Investigation of the Interaction between Epoxides and Collagen in Epoxy Tanning based on BDDGE Cross-linked Collagen Solution

2020 ◽  
Vol 115 (8) ◽  
pp. 279-287
Author(s):  
Yuanzhi Zhang ◽  
Guoying Li
Keyword(s):  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Decomposition Temperature ◽  
Diglycidyl Ether ◽  
Cross Linking ◽  
Force Microscopy ◽  
Atomic Force ◽  
Alkaline Conditions ◽  
The Cross ◽  
Dynamic Frequency

To investigate further the interaction between epoxides and collagen in epoxy tanning, collagen solutions (3 mg/ml) cross-linked with various concentrations (0.5–4-wt%) of 1,4-butanediol diglycidyl ether (BDDGE) at low temperature (4ºC), alkaline conditions (pH = 10) were prepared. The sodium dodecyl sulfate–polyacrylamide gel electrophoresis and Fourier transform infrared spectroscopy confirmed the intact triple-helix structure of the cross-linked collagen. With the increasing concentration of BDDGE, the denaturation temperature measured using VP-DSC increased from 42.56 ºC to 44.25ºC and thermogravimetric analysis showed that the decomposition temperature increased from 333.0ºC to 351.8ºC. In addition, the rheology properties such as G¢, G² and ?* were measured with a rotary rheometer using dynamic frequency scanning. The trinitrobenzensulfonic acid method and atomic force microscopy were used to investigate the interaction between collagen and BDDGE. The results indicated that the changes in cross-linked collagen performance were attributed to the transition of collagen aggregation caused by cross-linking. In addition, the transition point of 2-wt% BDDGE was the key node for the formation of a mature cross-linked network and the cross-linking barely increased above that. It is hoped that these findings deepen the understanding of epoxy tanning and provide guidance for the practical use of epoxides in tanning and biomaterials.

scholarly journals Chemical cross-linking of a dimeric protein on a modified lectin matrix. A general probe for the chemical topology of oligomeric glycoproteins

1981 ◽  
Vol 193 (3) ◽  
pp. 825-828 ◽  
Author(s):  
S Pillai
Keyword(s):  
Glucose Oxidase ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Cross Linking ◽  
Low Density ◽  
Amino Groups ◽  
Lysine Residues ◽  
The Cross ◽  
Interfacial Surfaces ◽  
Chemical Cross Linking

A dimeric glycoprotein, glucose oxidase, was allowed to react with lysine-specific cross-linkers, both when immobilized on a succinoylated lectin matrix at a critically low density and also at a high density in solution. Analysis of the cross-linked complexes thus obtained led to the following inferences with regard to the structure of this protein. (1) Of the 15 lysine residues on each glucose oxidase protomer, none is available on the non-interfacial surfaces. (2) Assuming that this protein possesses C2 symmetry with isologous bonding between subunits, it may be inferred that on each promoter there are at least two lysine clusters along or close to the interprotomeric interface. (3) These ‘interfacial’ lysine residues on each protomer are so oriented that the epsilon-amino groups of lysine residues a and b on protomer 1 ‘face’, and are very close to, the epsilon-amino groups of lysine residues b' and a' respectively on protomer 2. General inferences on the geometry of dimeric proteins derivable from an analysis of the cross-linked complexes obtained (as well as those not seen) by using this low-density matrix cross-linking approach were enumerated. Modified lectin matrices may prove useful in studying the three-dimensional structure of glycoproteins, particularly non-crystallizable oligomers.

Germin: physicochemical properties of the glycoprotein which signals onset of growth in the germinating wheat embryo

1987 ◽  
Vol 65 (12) ◽  
pp. 1039-1048 ◽  
Author(s):  
William C. McCubbin ◽  
Cyril M. Kay ◽  
Theresa D. Kennedy ◽  
Byron G. Lane
Keyword(s):  
Circular Dichroism ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Sedimentation Coefficient ◽  
Helical Structure ◽  
Random Coil ◽  
Sedimentation Equilibrium ◽  
Cross Linking ◽  
Wheat Embryo ◽  
Circular Dichroïsm

The size and structure of germin, the homooligomeric glycoprotein which marks the onset of growth in germinating wheat embryos, has been examined by gel filtration, ultracentrifugation, electron microscopy, chemical cross-linking, and optical techniques (circular dichroism). Germin has a sedimentation coefficient (S20,w) of 7.3S, and a Stokes' radius (RS) of 4.5 nm, the latter value being compatible with the dimensions of the particle observed by negative staining in the electron microscope. By three methods (sedimentation equilibrium, sodium dodecyl sulphate (SDS) – polyacrylamide electrophoresis, S20,w/RS), the mean particle mass of the two closely related forms of germin (G and G′) is ca. 130 kilodaltons (kDa). Cross-linking with dimethyl suberimidate indicates that the oligomer is homopentameric, compatible with the molecular mass of the protomer (ca. 26 kDa) as determined by SDS–polyacrylamide gel electrophoresis. Using the Provencher and Glockner analysis to interpret circular dichroism measurements (in the far ultraviolet), both forms of germin contain about 10–20% α-helical structure, 50–60% β-sheet/turn structure, and 20–30% random coil. In a structure-inducing environment (45% trifluoroethanol), the α-helical structure increases to a value (35–40%) similar to that predicted by Chou–Fasman analysis of the protein sequence deduced by cDNA sequencing.

scholarly journals p-NN′-phenylenebismaleimide, a specific cross-linking agent for F-actin

1978 ◽  
Vol 175 (3) ◽  
pp. 1023-1032 ◽  
Author(s):  
P Knight ◽  
G Offer
Keyword(s):  
Quantitative Analysis ◽  
Sodium Dodecyl Sulphate ◽  
Actin Filaments ◽  
Sodium Dodecyl ◽  
Cysteine Residue ◽  
Cross Linking ◽  
Maximum Value ◽  
Cross Links ◽  
The Cross ◽  
Hydrolysis Of

Covalent cross-links can be inserted between the subunits of F-actin by using p-NN′-phenylenebismaleimide. Cross-linking reaches its maximum value when one molecule of reagent has reacted with each actin subunit. p-NN′-Phenylenebismaleimide reacts initially with a cysteine residue on one subunit, the slower cross-linking reaction involving a lysine residue on a neighbouring subunit. Hydrolysis of the actin-bound reagent limits the extent of cross-linking. Quantitative analysis of the amounts of cross-linked oligomers seen on polyacrylamide gels containing sodium dodecyl sulphate suggests that neither the binding of the reagent to actin nor the formation of cross-links introduces strain into the structure. The cross-links do not join together different F-actin filaments, and evidence is presented that suggests that the cross-links join subunits of the same long-pitched helix.

scholarly journals Nanocellulose-Based Inks—Effect of Alginate Content on the Water Absorption of 3D Printed Constructs

2019 ◽  
Vol 6 (3) ◽  
pp. 65 ◽  
Author(s):  
Eduardo Espinosa ◽  
Daniel Filgueira ◽  
Alejandro Rodríguez ◽  
Gary Chinga-Carrasco
Keyword(s):  
Water Absorption ◽  
Moisture Absorption ◽  
Cellulose Nanofibrils ◽  
Three Dimensional ◽  
Wound Dressings ◽  
Cross Linking ◽  
Porous Structures ◽  
Freeze Dried ◽  
3D Printed ◽  
The Cross

2,2,6,6-tetramethylpyperidine-1-oxyl (TEMPO) oxidized cellulose nanofibrils (CNF) were used as ink for three-dimensional (3D) printing of porous structures with potential as wound dressings. Alginate (10, 20, 30 and 40 wt%) was incorporated into the formulation to facilitate the ionic cross-linking with calcium chloride (CaCl2). The effect of two different concentrations of CaCl2 (50 and 100 mM) was studied. The 3D printed hydrogels were freeze-dried to produce aerogels which were tested for water absorption. Scanning Electronic Microscopy (SEM) pictures demonstrated that the higher the concentration of the cross-linker the higher the definition of the printed tracks. CNF-based aerogels showed a remarkable water absorption capability. Although the incorporation of alginate and the cross-linking with CaCl2 led to shrinkage of the 3D printed constructs, the approach yielded suitable porous structures for water and moisture absorption. It is concluded that the 3D printed biocomposite structures developed in this study have characteristics that are promising for wound dressings devices.

Substitution of the γ-chain Asn308 disturbs the D:D interface affecting fibrin polymerization, fibrinopeptide B release, and FXIIIa-catalyzed cross-linking

Blood ◽  
2004 ◽  
Vol 103 (11) ◽  
pp. 4157-4163 ◽  
Author(s):  
Nobuo Okumura ◽  
Oleg V. Gorkun ◽  
Fumiko Terasawa ◽  
Susan T. Lord
Keyword(s):  
High Performance ◽  
Polyacrylamide Gel Electrophoresis ◽  
Sodium Dodecyl ◽  
Factor Xiiia ◽  
Cross Linking ◽  
Dimer Formation ◽  
Polymer Formation ◽  
Crystallographic Structures ◽  
Link Formation

Abstract Crystallographic structures indicate that γ-chain residue Asn308 participates in D:D interactions and indeed substitutions of γAsn308 with lysine or isoleucine have been identified in dysfibrinogens with impaired polymerization. To probe the role of Asn308 in polymerization, we synthesized 3 variant fibrinogens: γAsn308 changed to lysine (γN308K), isoleucine (γN308I), and alanine (γN308A). We measured thrombin-catalyzed polymerization by turbidity, fibrinopeptide release by high-performance liquid chromatography, and factor XIIIa–catalyzed cross-linking by sodium dodecyl sulfate–polyacrylamide gel electrophoresis. In the absence of added calcium, polymerization was clearly impaired with all 3 variants. In contrast, at 0.1 mM calcium, only polymerization of γN308K remained markedly abnormal. The release of thrombin-catalyzed fibrinopeptide B (FpB) was delayed in the absence of calcium, whereas at 1 mM calcium FpB release was delayed only with γN308K. Factor XIIIa–catalyzed γ-γ dimer formation was delayed with fibrinogen (in absence of thrombin), whereas with fibrin (in presence of thrombin) γ-γ dimer formation of only γN308K was delayed. These data corroborate the recognized link between FpB release and polymerization. They show fibrin cross-link formation likely depends on the structure of protofibrils. Together, our results show substitution of Asn308 with a hydrophobic residue altered neither polymer formation nor polymer structure at physiologic calcium concentrations, whereas substitution with lysine altered both.

scholarly journals Obtaining and Characterizing Alginate/k-Carrageenan Hydrogel Cross-Linked with Adipic Dihydrazide

2013 ◽  
Vol 2013 ◽  
pp. 1-12 ◽  
Author(s):  
Violeta Paşcalău ◽  
Violeta Popescu ◽  
George L. Popescu ◽  
Mircea C. Dudescu ◽  
Gheorghe Borodi ◽  
...  
Keyword(s):  
Cross Linking ◽  
Sodium Metaperiodate ◽  
Second Stage ◽  
End Groups ◽  
Reactive Compound ◽  
The Cross ◽  
Chemical Cross Linking

The aim of this paper is obtaining and characterizing hydrogels based on different ratios of oxidized alginate (oA) and k-carrageenan (C), chemically cross-linked with adipic dihydrazide (adh). The alginate (A) was first oxidized with sodium metaperiodate in order to transform it into the dialdehyde derivative, a more reactive compound than alginate. A known procedure for oxidation of alginate with sodium metaperiodate in ethanol-water in order to improve alginate reactivity by transforming the hydroxyl end-groups into dialdehyde was used, preceded by a partially cleavage of the alginate chains. In the second stage, the mixture of dialdehydic derivative of oxidized alginate, k-carrageenan and glycerol subjected to reaction with adipic dihydrazide leads to a Semi-Interpenetrated Network covalently cross-linked alginate/k-carrageenan hydrogel (oACadh), based on the dihydrazone compound which is responsible for the chemical cross-linking. Pure alginate, k-carrageenan, oxidized alginate, adipic dihydrazide and the cross-linked hydrogel were characterized by: FTIR, XRD, and SEM.

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