Direct NMR Detection of the Binding of Functional Ligands to the Human Sweet Receptor, a Heterodimeric Family 3 GPCR

Assadi‐Porter, Fariba M.; Tonelli, Marco; Maillet, Emeline; Hallenga, Klaas; Benard, Outhiriaradjou; Max, Marianna; Markley, John L.

doi:10.1021/ja8016939

Cited by 69 publications

(65 citation statements)

References 25 publications

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“…In addition, we are currently extending our studies to other envelope proteins including those of Ebola, influenza and SARS coronavirus. Importantly, the technique of STD NMR may in principle be applied to study of even larger systems such as whole virus or cells (31,32).…”

Section: Discussionmentioning

confidence: 99%

Probing the HIV gp120 Envelope Glycoprotein Conformation by NMR

Celigoy

Ramirez

Lin

et al. 2011

Journal of Biological Chemistry

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The HIV envelope glycoprotein gp120 plays a critical role in virus entry, and thus, its structure is of extreme interest for the development of novel therapeutics and vaccines. To date, high resolution structural information about gp120 in complex with gp41 has proven intractable. In this study, we characterize the structural properties of gp120 in the presence and absence of gp41 domains by NMR. Using the peptide probe 12p1 (sequence, RINNIPWSEAMM), which was identified previously as an entry inhibitor that binds to gp120, we identify atoms of 12p1 in close contact with gp120 in the monomeric and trimeric states. Interestingly, the binding mode of 12p1 with gp120 is similar for clades B and C. In addition, we show a subtle difference in the binding mode of 12p1 in the presence of gp41 domains, i.e. the trimeric state, which we interpret as small differences in the gp120 structure in the presence of gp41.

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Section: Discussionmentioning

confidence: 99%

Probing the HIV gp120 Envelope Glycoprotein Conformation by NMR

Celigoy

Ramirez

Lin

et al. 2011

Journal of Biological Chemistry

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“…S2B). Because thioredoxin has been used as a stabilizing partner of LysM1-2, we optimized the STD conditions to perform saturation transfer double-difference (STDD) NMR experiments useful to overcome strong background signals and to subtract possible artifacts because of contingent unspecific interactions between ligands and the stabilizing partner (22)(23)(24). The STDD NMR requires the acquisition of an additional STD spectrum of the receptor in absence of ligands, which is then subtracted from the STD spectrum acquired in the presence of ligands.…”

Section: Std Nmr Epitope Mapping Of Chitin Oligosaccharides In the Prmentioning

confidence: 99%

Chitin-induced activation of immune signaling by the rice receptor CEBiP relies on a unique sandwich-type dimerization

Hayafune

Berisio

Marchetti

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

284

250

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Perception of microbe-associated molecular patterns (MAMPs) through pattern recognition receptors (PRRs) triggers various defense responses in plants. This MAMP-triggered immunity plays a major role in the plant resistance against various pathogens. To clarify the molecular basis of the specific recognition of chitin oligosaccharides by the rice PRR, CEBiP (chitin-elicitor binding protein), as well as the formation and activation of the receptor complex, biochemical, NMR spectroscopic, and computational studies were performed. Deletion and domain-swapping experiments showed that the central lysine motif in the ectodomain of CEBiP is essential for the binding of chitin oligosaccharides. Epitope mapping by NMR spectroscopy indicated the preferential binding of longer-chain chitin oligosaccharides, such as heptamer-octamer, to CEBiP, and also the importance of N-acetyl groups for the binding. Molecular modeling/docking studies clarified the molecular interaction between CEBiP and chitin oligosaccharides and indicated the importance of Ile 122 in the central lysine motif region for ligand binding, a notion supported by site-directed mutagenesis. Based on these results, it was indicated that two CEBiP molecules simultaneously bind to one chitin oligosaccharide from the opposite side, resulting in the dimerization of CEBiP. The model was further supported by the observations that the addition of (GlcNAc) 8 induced dimerization of the ectodomain of CEBiP in vitro, and the dimerization and (GlcNAc) 8 -induced reactive oxygen generation were also inhibited by a unique oligosaccharide, (GlcNβ1,4GlcNAc) 4 , which is supposed to have N-acetyl groups only on one side of the molecule. Based on these observations, we proposed a hypothetical model for the ligand-induced activation of a receptor complex, involving both CEBiP and Oryza sativa chitinelicitor receptor kinase-1.plant immunity | MTI/PTI | chitin signaling | receptor-ligand interaction |

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“…The ligand-based methods, which typically involve comparison of the NMR parameters of a mixture of compounds in the presence and absence of the protein molecules, rely on the exchange-mediated transfer of bound state information to the free state. This biases ligand-based methods towards identification of weakly binding ligands (rapid exchange), but it does render the molecular weight of the protein molecule largely irrelevant, and indeed the approach has been applied to the ribosome, to receptors in membranes and to whole cells [64][65][66][67][68][69][70]. Competition experiments can be used to extend the range of the relaxation approach to tighter binding ligands, including those which are not in fast exchange.…”

Section: Detecting Binding -Nmr Screeningmentioning

confidence: 99%

Structural and Dynamic Information on Ligand Binding

Roberts¹

2011

Protein NMR Spectroscopy: Practical Techniques and Applications

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The function of the vast majority of proteins in the cell depends upon their noncovalent interactions with other molecules, ranging from enzyme-substrate interactions to the array of protein-protein and protein-nucleic acid interactions involved in, for example, the regulation of transcription. NMR has proved very valuable in the study of the interaction of proteins with both small and large molecules. In this chapter the focus will be on the basic principles of the study of ligand binding by NMR and on applications to small moleculeprotein interactions. Studies of macromolecular complexes are covered in Chapter 8. The use of NMR to study protein-ligand interactions is a vast area and, for reasons of space, many of the references herein are to reviews rather than to original papers; my apologies to those whose important contributions have not been cited directly.NMR can of course provide information on the structure of protein-ligand complexes, just as it can on the structure of the protein itself. However, the NMR spectrum is sensitive to both the equilibrium and the kinetics of the binding process. These effects can provide valuable information in themselves but, most importantly, they must be understood in order to interpret the spectrum correctly.

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Direct NMR Detection of the Binding of Functional Ligands to the Human Sweet Receptor, a Heterodimeric Family 3 GPCR

Cited by 69 publications

References 25 publications

Probing the HIV gp120 Envelope Glycoprotein Conformation by NMR

Probing the HIV gp120 Envelope Glycoprotein Conformation by NMR

Chitin-induced activation of immune signaling by the rice receptor CEBiP relies on a unique sandwich-type dimerization

Structural and Dynamic Information on Ligand Binding

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