scholarly journals Crystal structure of EndoS, an immunomodulatory endoglycosidase specific for human IgG antibodies

2014 ◽  
Vol 70 (a1) ◽  
pp. C256-C256
Author(s):  
Beatriz Trastoy
Keyword(s):  
Crystal Structure ◽  
Conformational Changes ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
Igg Antibodies ◽  
Human Pathogens ◽  
X Ray ◽  
Effector Functions ◽  
Encounter Complex ◽  
Key Enzyme

In order to evade host immune mechanisms, many bacteria secrete immunomodulatory enzymes. Streptococcus pyogenes, one of the most common human pathogens, secretes a large endoglycosidase, EndoS, which removes carbohydrates in a highly specific manner from IgG antibodies. This modification renders antibodies incapable of eliciting host effector functions through either complement or Fc γ receptors, providing the bacteria with a survival advantage. On account of this antibody-specific modifying activity, EndoS is being developed as a promising injectable therapeutic for autoimmune diseases that rely on autoantibodies. Additionally, EndoS is a key enzyme used in the chemoenzymatic synthesis of homogenously glycosylated antibodies with tailored Fc γ receptor-mediated effector functions. Despite the tremendous utility of this enzyme, the molecular basis of EndoS specificity for, and processing of, IgG antibodies has remained poorly understood. Here, we report the X-ray crystal structure of EndoS and provide a model of its encounter complex with its substrate, the IgG1 Fc domain. We show that EndoS is composed of five distinct protein domains, including glycosidase, leucine-rich repeat, hybrid Ig, carbohydrate binding module, and three-helix bundle domains, arranged in a distinctive V-shaped conformation. Our data suggest that the substrate enters the concave interior of the enzyme structure, is held in place by the carbohydrate binding module, and that concerted conformational changes in both enzyme and substrate are required for subsequent antibody deglycosylation. The EndoS structure presented here provides a framework from which novel endoglycosidases could be engineered for additional clinical and biotechnological applications.

X-ray Crystal Structure of a Non-crystalline Cellulose-specific Carbohydrate-binding Module: CBM28

2004 ◽  
Vol 339 (2) ◽  
pp. 253-258 ◽  
Author(s):  
Sheelan Jamal ◽  
Didier Nurizzo ◽  
Alisdair B. Boraston ◽  
Gideon J. Davies
Keyword(s):  
Crystal Structure ◽  
Crystalline Cellulose ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
X Ray

Comparative Analysis of Crystallinity Changes in Cellulose I Polymers Using ATR-FTIR, X-ray Diffraction, and Carbohydrate-Binding Module Probes

2011 ◽  
Vol 12 (11) ◽  
pp. 4121-4126 ◽  
Author(s):  
Alenka Kljun ◽  
Thomas A. S. Benians ◽  
Florence Goubet ◽  
Frank Meulewaeter ◽  
J. Paul Knox ◽  
...  
Keyword(s):  
Comparative Analysis ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
X Ray Diffraction ◽  
Cellulose I ◽  
X Ray ◽  
Crystallinity Changes

Structural basis of the transactivation deficiency of the human PPARγ F360L mutant associated with familial partial lipodystrophy

2014 ◽  
Vol 70 (7) ◽  
pp. 1965-1976 ◽  
Author(s):  
Clorinda Lori ◽  
Alessandra Pasquo ◽  
Roberta Montanari ◽  
Davide Capelli ◽  
Valerio Consalvi ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Conformational Changes ◽  
Partial Agonist ◽  
Salt Bridge ◽  
Van Der Waals Interactions ◽  
Structural Basis ◽  
Wild Type ◽  
Partial Lipodystrophy ◽  
X Ray ◽  
Familial Partial Lipodystrophy

The peroxisome proliferator-activated receptors (PPARs) are transcription factors that regulate glucose and lipid metabolism. The role of PPARs in several chronic diseases such as type 2 diabetes, obesity and atherosclerosis is well known and, for this reason, they are the targets of antidiabetic and hypolipidaemic drugs. In the last decade, some rare mutations in human PPARγ that might be associated with partial lipodystrophy, dyslipidaemia, insulin resistance and colon cancer have emerged. In particular, the F360L mutant of PPARγ (PPARγ2 residue 388), which is associated with familial partial lipodystrophy, significantly decreases basal transcriptional activity and impairs stimulation by synthetic ligands. To date, the structural reason for this defective behaviour is unclear. Therefore, the crystal structure of PPARγ F360L together with the partial agonist LT175 has been solved and the mutant has been characterized by circular-dichroism spectroscopy (CD) in order to compare its thermal stability with that of the wild-type receptor. The X-ray analysis showed that the mutation induces dramatic conformational changes in the C-terminal part of the receptor ligand-binding domain (LBD) owing to the loss of van der Waals interactions made by the Phe360 residue in the wild type and an important salt bridge made by Arg357, with consequent rearrangement of loop 11/12 and the activation function helix 12 (H12). The increased mobility of H12 makes the binding of co-activators in the hydrophobic cleft less efficient, thereby markedly lowering the transactivation activity. The spectroscopic analysis in solution and molecular-dynamics (MD) simulations provided results which were in agreement and consistent with the mutant conformational changes observed by X-ray analysis. Moreover, to evaluate the importance of the salt bridge made by Arg357, the crystal structure of the PPARγ R357A mutant in complex with the agonist rosiglitazone has been solved.

scholarly journals Domain structure of human complement C4b extends with increasing NaCl concentration: implications for its regulatory mechanism

2016 ◽  
Vol 473 (23) ◽  
pp. 4473-4491 ◽  
Author(s):  
Ka Wai Fung ◽  
David W. Wright ◽  
Jayesh Gor ◽  
Marcus J. Swann ◽  
Stephen J. Perkins
Keyword(s):  
Crystal Structure ◽  
Domain Structure ◽  
Conformational Changes ◽  
Analytical Ultracentrifugation ◽  
Sedimentation Coefficient ◽  
Salt Bridge ◽  
X Ray ◽  
X Ray Scattering ◽  
Nacl Concentration ◽  
Dual Polarisation Interferometry

During the activation of complement C4 to C4b, the exposure of its thioester domain (TED) is crucial for the attachment of C4b to activator surfaces. In the C4b crystal structure, TED forms an Arg104–Glu1032 salt bridge to tether its neighbouring macroglobulin (MG1) domain. Here, we examined the C4b domain structure to test whether this salt bridge affects its conformation. Dual polarisation interferometry of C4b immobilised at a sensor surface showed that the maximum thickness of C4b increased by 0.46 nm with an increase in NaCl concentration from 50 to 175 mM NaCl. Analytical ultracentrifugation showed that the sedimentation coefficient s20,w of monomeric C4b of 8.41 S in 50 mM NaCl buffer decreased to 7.98 S in 137 mM NaCl buffer, indicating that C4b became more extended. Small angle X-ray scattering reported similar RG values of 4.89–4.90 nm for C4b in 137–250 mM NaCl. Atomistic scattering modelling of the C4b conformation showed that TED and the MG1 domain were separated by 4.7 nm in 137–250 mM NaCl and this is greater than that of 4.0 nm in the C4b crystal structure. Our data reveal that in low NaCl concentrations, both at surfaces and in solution, C4b forms compact TED–MG1 structures. In solution, physiologically relevant NaCl concentrations lead to the separation of the TED and MG1 domain, making C4b less capable of binding to its complement regulators. These conformational changes are similar to those seen previously for complement C3b, confirming the importance of this salt bridge for regulating both C4b and C3b.

scholarly journals The first crystal structure of a family 31 carbohydrate-binding module with affinity to β-1,3-xylan

FEBS Letters ◽  
2005 ◽  
Vol 579 (20) ◽  
pp. 4324-4328 ◽  
Author(s):  
Hiroshi Hashimoto ◽  
Youichi Tamai ◽  
Fumiyoshi Okazaki ◽  
Yutaka Tamaru ◽  
Toshiyuki Shimizu ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Module

scholarly journals Crystal structure of recombinant tyrosinase-binding protein MtaL at 1.35 Å resolution

2016 ◽  
Vol 72 (3) ◽  
pp. 244-250 ◽  
Author(s):  
Xuelei Lai ◽  
Montserrat Soler-Lopez ◽  
Wangsa T. Ismaya ◽  
Harry J. Wichers ◽  
Bauke W. Dijkstra
Keyword(s):  
Crystal Structure ◽  
Conformational Changes ◽  
Agaricus Bisporus ◽  
Physiological Function ◽  
Carbohydrate Binding ◽  
Mushroom Tyrosinase ◽  
Binding Interface ◽  
Loop Regions ◽  
Carbohydrate Binding Site

Mushroom tyrosinase-associated lectin-like protein (MtaL) binds to matureAgaricus bisporustyrosinasein vivo, but the exact physiological function of MtaL is unknown. In this study, the crystal structure of recombinant MtaL is reported at 1.35 Å resolution. Comparison of its structure with that of the truncated and cleaved MtaL present in the complex with tyrosinase directly isolated from mushroom shows that the general β-trefoil fold is conserved. However, differences are detected in the loop regions, particularly in the β2–β3 loop, which is intact and not cleaved in the recombinant MtaL. Furthermore, the N-terminal tail is rotated inwards, covering the tyrosinase-binding interface. Thus, MtaL must undergo conformational changes in order to bind mature mushroom tyrosinase. Very interestingly, the β-trefoil fold has been identified to be essential for carbohydrate interaction in other lectin-like proteins. Comparison of the structures of MtaL and a ricin-B-like lectin with a bound disaccharide shows that MtaL may have a similar carbohydrate-binding site that might be involved in glycoreceptor activity.

scholarly journals Crystal structure of rhodopsin bound to arrestin by femtosecond X-ray laser

2014 ◽  
Vol 70 (a1) ◽  
pp. C567-C567
Author(s):  
H. Eric Xu
Keyword(s):  
Crystal Structure ◽  
G Protein ◽  
Conformational Changes ◽  
Molten Globule ◽  
Active Form ◽  
New Paradigm ◽  
X Ray ◽  
Biased Signaling ◽  
G Protein Coupled ◽  
Β Sheet

G protein-coupled receptors (GPCRs) signal primarily through G proteins or arrestins. Arrestin binding to GPCRs blocks G protein interaction and redirects signaling to numerous G protein-independent pathways. One structure of a GPCR bound to a G protein was solved, but the structure of a GPCR-arrestin complex has remained unknown despite its central role in GPCR biology. Here we report the crystal structure of a constitutively active form of human rhodopsin bound to a pre-activated form of the mouse visual arrestin, determined by serial femtosecond X-ray laser crystallography. The structure reveals that arrestin binding induces large and unexpected conformational changes at both the extracellular and intracellular sides of rhodopsin. Arrestin also undergoes dramatic rearrangements from its inactive well-ordered β-sheet structure into a more flexible molten globule-type state, allowing a snake-like movement of the first 77 arrestin residues that shortens its central crest finger loop by seven residues to accommodate the concave surface of rhodopsin. This structure provides a basis for understanding GPCR-mediated arrestin-biased signaling, reveals a new paradigm of signal transduction by a molten globule, and demonstrates the extraordinary power of X-ray lasers for advancing the frontiers of structural biology.

scholarly journals Crystallization and preliminary X-ray diffraction studies of the family 54 carbohydrate-binding module from laminarinase (β-1,3-glucanase) Lic16A ofClostridium thermocellum

2015 ◽  
Vol 71 (2) ◽  
pp. 217-220 ◽  
Author(s):  
Yury A. Kislitsyn ◽  
Valeriya R. Samygina ◽  
Igor A. Dvortsov ◽  
Nataliya A. Lunina ◽  
Inna P. Kuranova ◽  
...  
Keyword(s):  
Three Dimensional ◽  
Dimensional Structure ◽  
Diffusion Method ◽  
Carbohydrate Binding ◽  
Carbohydrate Binding Module ◽  
X Ray Diffraction ◽  
X Ray ◽  
Cell Parameters ◽  
The Family ◽  
Overexpression System

The crystallization and preliminary X-ray diffraction analysis of the carbohydrate-binding module (CBM) from laminarinase Lic16A of the hyperthermophilic anaerobic bacteriumClostridium thermocellum(ctCBM54) are reported. Recombinant ctCBM54 was prepared using anEscherichia coli/pQE30 overexpression system and was crystallized by the hanging-drop vapour-diffusion method. X-ray diffraction data were collected to 2.1 Å resolution using synchrotron radiation. The crystals belonged to space groupP6322, with unit-cell parametersa=b= 130.15,c= 131.05 Å. The three-dimensional structure of ctCBM54 will provide valuable information about the structure–function relation of the laminarinase Lic16A and will allow the exploitation of this binding module in biotechnological applications.

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