An unusual antithrombin-binding heparin octasaccharide with an additional 3-O-sulfated glucosamine in the active pentasaccharide sequence

2012 ◽  
Vol 449 (2) ◽  
pp. 343-351 ◽  
Author(s):  
Marco Guerrini ◽  
Stefano Elli ◽  
Pierre Mourier ◽  
Timothy R. Rudd ◽  
Davide Gaudesi ◽  
...  
Keyword(s):  
Molecular Conformation ◽  
Biochemical Characterization ◽  
Saturation Transfer ◽  
Antithrombotic Activity ◽  
Saturation Transfer Difference ◽  
Order Of Magnitude ◽  
Epitope Binding ◽  
Docking Energy ◽  
Magnitude Increase

The 3-O-sulfation of N-sulfated glucosamine is the last event in the biosynthesis of heparin/heparan sulfate, giving rise to the antithrombin-binding pentasaccharide sequence AGA*IA, which is largely associated with the antithrombotic activity of these molecules. The aim of the present study was the structural and biochemical characterization of a previously unreported AGA*IA*-containing octasaccharide isolated from the very-low-molecular-mass heparin semuloparin, in which both glucosamine residues of the pentasaccharide moiety located at the non-reducing end bear 3-O-sulfate groups. Two-dimensional and STD (saturation transfer difference) NMR experiments clearly confirmed its structure and identified its ligand epitope binding to antithrombin. The molecular conformation of the octasaccharide–antithrombin complex has been determined by NMR experiments and docking/energy minimization. The presence of the second 3-O-sulfated glucosamine in the octasaccharide induced more than one order of magnitude increase in affinity to antithrombin compared to the pentasaccharide AGA*IA.

scholarly journals Biochemical characterization of the catalytic domains of three different clostridial collagenases

2009 ◽  
Vol 390 (1) ◽  
Author(s):  
Ulrich Eckhard ◽  
Esther Schönauer ◽  
Paulina Ducka ◽  
Peter Briza ◽  
Dorota Nüss ◽  
...  
Keyword(s):  
Biochemical Characterization ◽  
Substrate Affinity ◽  
Catalytic Domains ◽  
Zinc Binding Site ◽  
N Terminus ◽  
Activation Mechanisms ◽  
Order Of Magnitude ◽  
Therapy And Diagnosis ◽  
Prime Target

Abstract Clostridial collagenases are used for a broad spectrum of biotechnological applications and represent prime target candidates for both therapy and diagnosis of clostridial infections. In this study, we biochemically characterized the catalytic domains of three clostridial collagenases, collagenase G (ColG) and H (ColH) from Clostridium histolyticum, and collagenase T (ColT) from C. tetani. All protein samples showed activity against a synthetic peptidic substrate (furylacryloyl-Leu-Gly-Pro-Ala, FALGPA) with ColH showing the highest overall activity and highest substrate affinity. Whereas the K m values of all three enzymes were within the same order of magnitude, the turnover rate k cat of ColG decreased 50- to 150-fold when compared to ColT and ColH. It is noteworthy that the protein N-terminus significantly impacts their substrate affinity and substrate turnover as well as their inhibition profile with 1,10-phenanthroline. These findings were complemented with the discovery of a strictly conserved double-glycine motif, positioned 28 amino acids upstream of the HEXXH zinc binding site, which is critical for enzymatic activity. These observations have consequences with respect to the topology of the N-terminus relative to the active site as well as possible activation mechanisms.

Saturation Transfer Difference (STD) NMR Spectroscopy Characterization of Dual Binding Mode of a Mannose Disaccharide to DC-SIGN

ChemBioChem ◽  
2008 ◽  
Vol 9 (14) ◽  
pp. 2225-2227 ◽  
Author(s):  
Jesús Angulo ◽  
Irene Díaz ◽  
José J. Reina ◽  
Georges Tabarani ◽  
Franck Fieschi ◽  
...  
Keyword(s):  
Nmr Spectroscopy ◽  
Binding Mode ◽  
Saturation Transfer ◽  
Saturation Transfer Difference ◽  
Std Nmr

scholarly journals Investigating the interaction mechanism of fluorescent whitening agents to human serum albumin using saturation transfer difference-NMR, multi-spectroscopy, and docking studies

RSC Advances ◽  
2017 ◽  
Vol 7 (44) ◽  
pp. 27796-27806 ◽  
Author(s):  
Ludan Zhao ◽  
Jiuyang Liu ◽  
Ronghui Guo ◽  
Qiaomei Sun ◽  
Hongqin Yang ◽  
...  
Keyword(s):  
Human Serum Albumin ◽  
Serum Albumin ◽  
Human Serum ◽  
Interaction Mechanism ◽  
Docking Studies ◽  
Saturation Transfer ◽  
Saturation Transfer Difference ◽  
Fluorescent Whitening Agents ◽  
Std Nmr

Characterization of the interaction between two fluorescent whitening agents and human serum albumin: 1H STD-NMR, multi-spectroscopy, and docking studies.

Characterization of heparin–protein interaction by saturation transfer difference (STD) NMR

2014 ◽  
Vol 406 (13) ◽  
pp. 3079-3089 ◽  
Author(s):  
Fei Yu ◽  
Sucharita Roy ◽  
Enrique Arevalo ◽  
John Schaeck ◽  
Jason Wang ◽  
...  
Keyword(s):  
Protein Interaction ◽  
Saturation Transfer ◽  
Saturation Transfer Difference ◽  
Std Nmr

scholarly journals Multifrequency STD NMR Unveils the Interactions of Antibiotics With Burkholderia multivorans Biofilm Exopolysaccharide

2021 ◽  
Vol 8 ◽  
Author(s):  
Ridvan Nepravishta ◽  
Serena Monaco ◽  
Marco Distefano ◽  
Roberto Rizzo ◽  
Paola Cescutti ◽  
...  
Keyword(s):  
Active Role ◽  
Atomic Level ◽  
Bacterial Cells ◽  
Saturation Transfer ◽  
Burkholderia Multivorans ◽  
Powerful Technique ◽  
Macromolecular Assemblies ◽  
Saturation Transfer Difference ◽  
Std Nmr

Biofilms confine bacterial cells within self-produced matrices, offering advantages such as protection from antibiotics and entrapment of nutrients. Polysaccharides are major components in these macromolecular assemblies, and their interactions with other chemicals are of high relevance for the benefits provided by the biofilm 3D molecular matrix. NMR is a powerful technique for the study and characterization of the interactions between molecules of biological relevance. In this study, we have applied multifrequency saturation transfer difference (STD) NMR and DOSY NMR approaches to elucidate the interactions between the exopolysaccharide produced by Burkholderia multivorans C1576 (EpolC1576) and the antibiotics kanamycin and ceftadizime. The NMR strategies presented here allowed for an extensive characterization at an atomic level of the mechanisms behind the implication of the EpolC1576 in the recalcitrance phenomena, which is the ability of bacteria in biofilms to survive in the presence of antibiotics. Our results suggest an active role for EpolC1576 in the recalcitrance mechanisms toward kanamycin and ceftadizime, though through two different mechanisms.

Plant Pathology Diagnosed by X-Ray Microanalysis

1987 ◽  
Vol 45 ◽  
pp. 974-975
Author(s):  
J. H. Resau ◽  
N. Howell ◽  
S. H. Chang
Keyword(s):  
Electron Microscopy ◽  
Plant Pathology ◽  
Biochemical Characterization ◽  
Nutrient Deficiency ◽  
Green Leaf ◽  
Parasitic Infestation ◽  
X Ray

Spinach grown in Texas developed “yellow spotting” on the peripheral portions of the leaves. The exact cause of the discoloration could not be determined as there was no evidence of viral or parasitic infestation of the plants and biochemical characterization of the plants did not indicate any significant differences between the yellow and green leaf portions of the spinach. The present study was undertaken using electron microscopy (EM) to determine if a micro-nutrient deficiency was the cause for the discoloration.Green leaf spinach was collected from the field and sent by express mail to the EM laboratory. The yellow and equivalent green portions of the leaves were isolated and dried in a Denton evaporator at 10-5 Torr for 24 hrs. The leaf specimens were then examined using a JEOL 100 CX analytical microscope. TEM specimens were prepared according to the methods of Trump et al.

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