scholarly journals Chemical modification studies on Abrus agglutinin. Involvement of tryptophan residues in sugar binding

1984 ◽  
Vol 217 (3) ◽  
pp. 773-781 ◽  
Author(s):  
S R Patanjali ◽  
M J Swamy ◽  
V Anantharam ◽  
M I Khan ◽  
A Surolia
Keyword(s):  
Binding Activity ◽  
Carbohydrate Binding ◽  
Amino Acid Residues ◽  
Partial Protection ◽  
Abrus Precatorius ◽  
Biphasic Pattern ◽  
Saccharide Binding ◽  
Tryptophan Residues ◽  
Haemagglutinating Activity ◽  
Binding Lectin

The galactose-binding lectin from the seeds of the jequirity plant (Abrus precatorius) was subjected to various chemical modifications in order to detect the amino acid residues involved in its binding activity. Modification of lysine, tyrosine, arginine, histidine, glutamic acid and aspartic acid residues did not affect the carbohydrate-binding activity of the agglutinin. However, modification of tryptophan residues carried out in native and denaturing conditions with N-bromosuccinimide and 2-hydroxy-5-nitrobenzyl bromide led to a complete loss of its carbohydrate-binding activity. Under denaturing conditions 30 tryptophan residues/molecule were modified by both reagents, whereas only 16 and 18 residues/molecule were available for modification by N-bromosuccinimide and 2-hydroxy-5-nitrobenzyl bromide respectively under native conditions. The relative loss in haemagglutinating activity after the modification of tryptophan residues indicates that two residues/molecule are required for the carbohydrate-binding activity of the agglutinin. A partial protection was observed in the presence of saturating concentrations of lactose (0.15 M). The decrease in fluorescence intensity of Abrus agglutinin on modification of tryptophan residues is linear in the absence of lactose and shows a biphasic pattern in the presence of lactose, indicating that tryptophan residues go from a similar to a different molecular environment on saccharide binding. The secondary structure of the protein remains practically unchanged upon modification of tryptophan residues, as indicated by c.d. and immunodiffusion studies, confirming that the loss in activity is due to modification only.

Studies on the tryptophan residues of soybean agglutinin. Involvement in saccharide binding

1989 ◽  
Vol 9 (2) ◽  
pp. 189-198 ◽  
Author(s):  
Musti Joginadha Swamy ◽  
Avadhesha Surolia
Keyword(s):  
Active Site ◽  
Blue Shift ◽  
Binding Activity ◽  
Carbohydrate Binding ◽  
Side Chains ◽  
Soybean Agglutinin ◽  
Protein Fluorescence ◽  
Saccharide Binding ◽  
Tryptophan Residues ◽  
Tryptophan Side Chains

Modification of tryptophan side chains of soybean agglutinin (SBA) with N-bromosuccinimide results in a loss of the hemagglutinating and carbohydrate binding activities of the protein. One residue/subunit is probably essential for the binding activity. Modification leads to a large decrease in the fluorescene of the protein accompained by a blue shift. Iodide ion quenching of the protein fluorescence shows that saccharide binding results in a decreased accessibility of some of the tryptophan side chains. These results strongly point towards the involvement of tryptophan residues in the active site of SBA.

scholarly journals Production and characterization of monoclonal antibodies to β-galactoside-binding lectin of bovine heart muscle. Direct evidence that haemagglutinating activity is associated with a 13kDa protein

1984 ◽  
Vol 220 (1) ◽  
pp. 253-260 ◽  
Author(s):  
S R Carding ◽  
R Thorpe ◽  
R A Childs ◽  
M Spitz ◽  
T Feizi
Keyword(s):  
Heart Muscle ◽  
Direct Evidence ◽  
Solid Phase ◽  
Binding Activity ◽  
High Titre ◽  
Immunoglobulin M ◽  
Bovine Heart ◽  
Haemagglutinating Activity ◽  
Binding Lectin

With the aim of obtaining monospecific antibodies against the beta-galactoside-binding lectin of bovine heart muscle, spleen cells from Lou rats immunized with lectin were fused with the rat myeloma line Y3.Ag1.2.3. Two immunoglobulin M (IgM)-producing clones, designated NIBy 142-36/8 and NIBy 143-9/5, derived from separate fusions, were used to generate ascites containing high-titre binding activity against the 13kDa component in preparations of lectin. Direct evidence that haemagglutinating activity is associated with the 13kDa protein was obtained by the specific elution of 13kDa polypeptides with haemagglutinating activity from an immobilized antibody adsorbent. Solid-phase radiobinding assays and immunoblotting of isolated lectins and/or muscle homogenates confirmed the earlier indications with conventional antisera that the beta-galactoside-binding lectins of bovine, human and monkey muscle tissue are antigenically related.

Mannan-binding lectin (MBL) production from human plasma

2003 ◽  
Vol 31 (4) ◽  
pp. 758-762 ◽  
Author(s):  
I. Laursen
Keyword(s):  
Affinity Chromatography ◽  
Production Process ◽  
Binding Activity ◽  
Carbohydrate Binding ◽  
Substitution Therapy ◽  
Safety Study ◽  
Low Levels ◽  
Mannan Binding Lectin ◽  
Binding Lectin ◽  
Mbl Deficiency

Individuals with low levels of mannan-binding lectin (MBL) appear to be susceptible to infectious diseases. This suggests that substitution therapy with MBL might be a beneficial treatment of patients with MBL deficiency. A production process for an MBL product has been developed from a fraction II+III precipitate obtained by ethanol fractionation of plasma. The MBL process includes three chromatographic steps, where the first and key step is affinity chromatography on a cross-linked agarose matrix selecting for oligomeric, carbohydrate-binding MBL. The yield from the production process is about 25% of the plasma MBL content, and the purity is about 65%. The MBL product shows mannan-binding activity and complement-activating ability. A safety study has shown this plasma-derived MBL to be safe and well tolerated in adult MBL-deficient volunteers.

Genomic analysis of C-type lectins

2002 ◽  
Vol 69 ◽  
pp. 59-72 ◽  
Author(s):  
Kurt Drickamer ◽  
Andrew J. Fadden
Keyword(s):  
Biological Effects ◽  
Cell Signalling ◽  
Genomic Analysis ◽  
Binding Activity ◽  
Immunoglobulin Superfamily ◽  
Carbohydrate Binding ◽  
Carbohydrate Recognition ◽  
Calcium Dependent ◽  
Binding Domains ◽  
Carbohydrate Recognition Domains

Many biological effects of complex carbohydrates are mediated by lectins that contain discrete carbohydrate-recognition domains. At least seven structurally distinct families of carbohydrate-recognition domains are found in lectins that are involved in intracellular trafficking, cell adhesion, cell–cell signalling, glycoprotein turnover and innate immunity. Genome-wide analysis of potential carbohydrate-binding domains is now possible. Two classes of intracellular lectins involved in glycoprotein trafficking are present in yeast, model invertebrates and vertebrates, and two other classes are present in vertebrates only. At the cell surface, calcium-dependent (C-type) lectins and galectins are found in model invertebrates and vertebrates, but not in yeast; immunoglobulin superfamily (I-type) lectins are only found in vertebrates. The evolutionary appearance of different classes of sugar-binding protein modules parallels a development towards more complex oligosaccharides that provide increased opportunities for specific recognition phenomena. An overall picture of the lectins present in humans can now be proposed. Based on our knowledge of the structures of several of the C-type carbohydrate-recognition domains, it is possible to suggest ligand-binding activity that may be associated with novel C-type lectin-like domains identified in a systematic screen of the human genome. Further analysis of the sequences of proteins containing these domains can be used as a basis for proposing potential biological functions.

scholarly journals Biochemical and Initial Structural Characterization of the Monocot Chimeric Jacalin OsJAC1

2021 ◽  
Vol 22 (11) ◽  
pp. 5639
Author(s):  
Nikolai Huwa ◽  
Oliver H. Weiergräber ◽  
Christian Kirsch ◽  
Ulrich Schaffrath ◽  
Thomas Classen
Keyword(s):  
Physiological Role ◽  
Basic Function ◽  
Binding Activity ◽  
Carbohydrate Binding ◽  
Lectin Domain ◽  
Reducing Conditions ◽  
Transition Points ◽  
The Individual ◽  
High Yields ◽  
Dirigent Domain

The monocot chimeric jacalin OsJAC1 from Oryza sativa consists of a dirigent and a jacalin-related lectin domain. The corresponding gene is expressed in response to different abiotic and biotic stimuli. However, there is a lack of knowledge about the basic function of the individual domains and their contribution to the physiological role of the entire protein. In this study, we have established a heterologous expression in Escherichia coli with high yields for the full-length protein OsJAC1 as well as its individual domains. Our findings showed that the secondary structure of both domains is dominated by β-strand elements. Under reducing conditions, the native protein displayed clearly visible transition points of thermal unfolding at 59 and 85 °C, which could be attributed to the lectin and the dirigent domain, respectively. Our study identified a single carbohydrate-binding site for each domain with different specificities towards mannose and glucose (jacalin domain), and galactose moieties (dirigent domain), respectively. The recognition of different carbohydrates might explain the ability of OsJAC1 to respond to different abiotic and biotic factors. This is the first report of specific carbohydrate-binding activity of a DIR domain, shedding new light on its function in the context of this monocot chimeric jacalin.

New structural insights into anomeric carbohydrate recognition by frutalin: an α-d-galactose-binding lectin from breadfruit seeds

2019 ◽  
Vol 476 (1) ◽  
pp. 101-113 ◽  
Author(s):  
Antonio Eufrásio Vieira Neto ◽  
Felipe Domingos de Sousa ◽  
Humberto D'Muniz Pereira ◽  
Frederico Bruno Mendes Batista Moreno ◽  
Marcos Roberto Lourenzoni ◽  
...  
Keyword(s):  
Amino Acids ◽  
Biological Activities ◽  
3D Structure ◽  
Carbohydrate Binding ◽  
X Ray Crystallography ◽  
Multiple Binding Sites ◽  
Multiple Binding ◽  
New Perspective ◽  
Dynamics Simulations ◽  
Binding Lectin

Abstract Frutalin (FTL) is a multiple-binding lectin belonging to the jacalin-related lectin (JRL) family and derived from Artocarpus incisa (breadfruit) seeds. This lectin specifically recognizes and binds α-d-galactose. FTL has been successfully used in immunobiological research for the recognition of cancer-associated oligosaccharides. However, the molecular bases by which FTL promotes these specific activities remain poorly understood. Here, we report the whole 3D structure of FTL for the first time, as determined by X-ray crystallography. The obtained crystals diffracted to 1.81 Å (Apo-frutalin) and 1.65 Å (frutalin–d-Gal complex) of resolution. The lectin exhibits post-translational cleavage yielding an α- (133 amino acids) and β-chain (20 amino acids), presenting a homotetramer when in solution, with a typical JRL β-prism. The β-prism was composed of three 4-stranded β-sheets forming three antiparallel Greek key motifs. The carbohydrate-binding site (CBS) involved the N-terminus of the α-chain and was formed by four key residues: Gly25, Tyr146, Trp147 and Asp149. Together, these results were used in molecular dynamics simulations in aqueous solutions to shed light on the molecular basis of FTL-ligand binding. The simulations suggest that Thr-Ser-Ser-Asn (TSSN) peptide excision reduces the rigidity of the FTL CBS, increasing the number of interactions with ligands and resulting in multiple-binding sites and anomeric recognition of α-d-galactose sugar moieties. Our findings provide a new perspective to further elucidate the versatility of FTL in many biological activities.

scholarly journals Interaction of dirhenium(III) tryptophan complex compound with DNA and protein

2021 ◽  
pp. 35-40
Author(s):  
O.A. Holichenko ◽  
◽  
N.I. Shtemenko ◽  
A.A. Ovcharenko ◽  
A.V. Shtemenko
Keyword(s):  
Complex Compound ◽  
Blue Shift ◽  
Binding Activity ◽  
Significant Shift ◽  
Amino Acid Residues ◽  
Conjugated Systems ◽  
Quadruple Bond ◽  
G4 Dna ◽  
Main Protein ◽  
Dna Silencing

We report about the interactions of dirhenium(III) compound cis-[Re2(Trp)2Cl4(CH3CN)2]Cl2 (I) with bovine serum protein (BSA) and guanine (G4) quadruplexes DNA by UV-Vis titration. Addition of I to BSA led to the interaction between these compounds with binding constant 5.6103 M–1 and hyperchromism (20.9%) of the main protein absorption band (280 nm). These results support our assumption about formation of the additional conjugated systems during the process of interaction with BSA. Stabilization of the quadruple bonded rhenium(III) complex compound was shown in the presence of BSA (the rate of destruction was reduced), that may be explained by interaction between amino acid residues of BSA and quadruple bond of dirhenium(III) complex compound. In addition, we have obtained data about strong hyperchromism (up to 100%) and significant shift of the maximum of absorption (blue shift) towards UV (2–9 nm) and visible (22 nm) regions in the spectra of mixtures G4s and I, that, in our opinion, correlated with a conformational change in DNA and with formation of additional conjugated systems around quadruple bond of I. In a whole, our work confirms the strong binding activity of a cluster dirhenium(III) compound towards G4 quadruplexes, that exceed the binding activity to proteins and witness to preferential interactions of I with G4 DNA in a living cell. These results may be used in DNA "silencing technology" and "antisense therapy".

Fluorescent Properties of Firefly Luciferases and Their Complexes with Luciferin

2000 ◽  
Vol 20 (1) ◽  
pp. 21-30 ◽  
Author(s):  
Ekaterina I. Dementieva ◽  
Elena A. Fedorchuk ◽  
Lubov Yu. Brovko ◽  
Alexander P. Savitskii ◽  
Natalya N. Ugarova
Keyword(s):  
Energy Transfer ◽  
Tryptophan Fluorescence ◽  
Tryptophan Residue ◽  
Amino Acid Residues ◽  
Photinus Pyralis ◽  
Fluorescent Properties ◽  
Effective Energy Transfer ◽  
Tryptophan Residues ◽  
Luciola Mingrelica

Fluorescence of luciferases from Luciola mingrelica (single tryptophanresidue, Trp-419) and Photinus pyralis (two tryptophan residues, Trp-417,Trp-426) was studied. Analysis of quenching of tryptophan fluorescenceshowed that the tryptophan residue conserved in all luciferases is notaccessible for charged quenchers, which is explained by the presence ofpositively and negatively charged amino acid residues in the close vicinityto it. An effective energy transfer from tryptophan to luciferin wasobserved during quenching of tryptophan fluorescence of both luciferaseswith luciferin. From the data on the energy transfer, the distance betweenthe luciferin molecule and Trp-417 (419) in the luciferin–luciferasecomplex was calculated: 11–15 Å for P. pyralis and 12–17Å for L. mingrelica luciferases. The role of the conserved Trp residuein the catalysis is discussed.

scholarly journals Energetics of carbohydrate binding by a 14 kDa S-type mammalian lectin

1995 ◽  
Vol 308 (1) ◽  
pp. 237-241 ◽  
Author(s):  
R Ramkumar ◽  
A Surolia ◽  
S K Podder
Keyword(s):  
Binding Constants ◽  
Van Der Waals Interactions ◽  
Titration Calorimetry ◽  
Carbohydrate Binding ◽  
Glucose Binding ◽  
The Stability ◽  
L 14 ◽  
Derivatives Of ◽  
Binding Lectin ◽  
Site Binding

The thermodynamics of the binding of derivatives of galactose and lactose to a 14 kDa beta-galactoside-binding lectin (L-14) from sheep spleen has been studied in 10 nM phosphate/150 mM NaCl/10 mM beta-mercaptoethanol buffer, pH 7.4, and in the temperature range 285-300 K using titration calorimetry. The single-site binding constants of various sugars for the lectin were in the following order: N-acetyl-lactosamine thiodigalactoside > 4-methylumbelliferyl lactoside > lactose > 4-methylumbelliferyl alpha-D-galactoside > methyl-alpha-galactose > methyl-beta-galactose. Reactions were essentially enthalpically driven with the binding enthalpies ranging from -53.8 kJ/mol for thiodigalactoside at 301 K to -2.2 kJ/mol for galactose at 300 K, indicating that hydrogen-bonding and van der Waals interactions provide the major stabilization for these reactions. However, the binding of 4-methylumbelliferyl-alpha-D-galactose displays relatively favourable entropic contributions, indicating the existence of a non-polar site adjacent to the galactose-binding subsite. From the increments in the enthalpies for the binding of lactose, N-acetyl-lactosamine and thiodigalactoside relative to methyl-beta-galactose, the contribution of glucose binding in the subsite adjacent to that for galactose shows that glucose makes a major contribution to the stability of L-14 disaccharide complexes. Observation of enthalpy-entropy compensation for the recognition of saccharides such as lactose by L-14 and the absence of it for monosaccharides such as galactose, together with the lack of appreciable changes in the heat capacity (delta Cp), indicate that reorganization of water plays an important role in these reactions.

Definition of Essential Amino Acid Residues in the Recognition of a Peptide by a Mouse Monoclonal Antibody

1997 ◽  
Vol 52 (3-4) ◽  
pp. 274-278
Author(s):  
Laura Rosanó ◽  
Francesca Di Modugno ◽  
Giulia Romagnoli ◽  
Alberto Chersi
Keyword(s):  
Monoclonal Antibody ◽  
Amino Acid ◽  
Critical Role ◽  
Binding Activity ◽  
Mouse Monoclonal Antibody ◽  
Amino Acid Residues ◽  
Binding Data ◽  
Amino Acid Stretch ◽  
Definition Of ◽  
Key Residues

AbstractA mouse monoclonal antibody reacting in ELISA with a synthetic peptide representing a linear amino acid stretch of the protein antigen was tested on all overlap­ ping 5-mer to 9-mer fragments of the peptide, as prepared by multi-pin synthesis. Analysis of the binding data suggests that several residues in the peptide might be relatively unrelevant for recognition, while few others seem to play a critical role as key residues. On the basis of such observations, we attempted to reconstruct an alternative essential epitope by introducing multiple amino acid substitutions in the 9-mer peptide exhibiting the best binding activity, and then tested its ability to be recognized by the monoclonal antibody.

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