Related ArticlesAnalysis of SNARE complex/Synaptotagmin-1 Interactions by One-dimensional NMR Spectroscopy.
Biochemistry. 2013 Apr 25;
Authors: Zhou A, Brewer KD, Rizo J
Abstract
Neurotransmitter release depends critically on the Ca2+ sensor synaptotagmin-1 and the SNARE proteins syntaxin-1, synaptobrevin and SNAP-25, which mediate membrane fusion by forming tight SNARE complexes that bridge the synaptic vesicle and plasma membranes. Interactions between the SNARE complex and the two C2 domains of synaptotagmin-1 (the C2A and C2B domains) are believed to play a key role in coupling Ca2+ sensing to membrane fusion, but the nature of these interactions is unclear, in part because of a paucity of data obtained by quantitative biophysical methods. Here we have analyzed synaptotagmin-1/SNARE complex interactions by monitoring the decrease in the intensities of one-dimensional 13C-edited 1H-NMR spectra of 13C-labeled fragments of synaptotagmin-1 upon binding to unlabeled SNARE complex. Our results indicate that there is a primary binding mode between synaptotagmin-1 and the SNARE complex that involves a polybasic region in the C2B domain and has a submicromolar affinity. Our NMR data, combined with precipitation assays, show that there are additional SNARE complex/synaptotagmin-1 interactions that lead to aggregation and that involve in part two arginines at the bottom of the C2B domain. Overall, this study shows the importance of disentangling the contributions of different types of interactions to SNARE complex/synaptotagmin-1 binding, and illustrate the usefulness of one-dimensional NMR methods to analyze intricate protein interactions.
PMID: 23617808 [PubMed - as supplied by publisher]
Analysis of complex mixtures using high-resolution nuclear magnetic resonance spectroscopy and chemometrics
Analysis of complex mixtures using high-resolution nuclear magnetic resonance spectroscopy and chemometrics
Publication year: 2011
Source: Progress in Nuclear Magnetic Resonance Spectroscopy, In Press, Accepted Manuscript, Available online 12 May 2011</br>
James S., McKenzie , James A., Donarski , Julie C., Wilson , Adrian J., Charlton</br>
*Highlights:*? Analysis of complex mixtures using NMR spectroscopy. ? Use of chemometrics for interpretation of spectra. ? Review of sample handling approaches. ? Discussion of spectral processing methods. ? Discussion of supervised and...
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[NMR paper] Facile detection of protein-protein interactions by one-dimensional NMR spectroscopy.
Facile detection of protein-protein interactions by one-dimensional NMR spectroscopy.
Related Articles Facile detection of protein-protein interactions by one-dimensional NMR spectroscopy.
Biochemistry. 2003 Mar 18;42(10):2774-80
Authors: Araç D, Murphy T, Rizo J
Two methods for detecting protein-protein interactions in solution using one-dimensional (1D) NMR spectroscopy are described. Both methods rely on measurement of the intensity of the strongest methyl resonance (SMR), which for most proteins is observed at 0.8-0.9 ppm. The severe...
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[NMR paper] Three-dimensional structure of the FK506 binding protein/ascomycin complex in solutio
Three-dimensional structure of the FK506 binding protein/ascomycin complex in solution by heteronuclear three- and four-dimensional NMR.
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Biochemistry. 1993 Jan 26;32(3):754-65
Authors: Meadows RP, Nettesheim DG, Xu RX, Olejniczak ET, Petros AM, Holzman TF, Severin J, Gubbins E, Smith H, Fesik SW
A high-resolution three-dimensional solution structure of the FKBP/ascomycin complex has been...
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08-21-2010 11:53 PM
[NMR paper] Two-dimensional NMR study of a protein-DNA complex. lac repressor headpiece-operator
Two-dimensional NMR study of a protein-DNA complex. lac repressor headpiece-operator interaction.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--linkinghub.elsevier.com-ihub-images-PubMedLink.gif Related Articles Two-dimensional NMR study of a protein-DNA complex. lac repressor headpiece-operator interaction.
Biochem Pharmacol. 1990 Jul 1;40(1):89-96
Authors: Kaptein R, Lamerichs RM, Boelens R, Rullmann JA
The interaction of the N-terminal DNA-binding domain (56 amino acid residues) of the lac repressor with lac operator DNA was...
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[NMR paper] The three-dimensional structure of the vnd/NK-2 homeodomain-DNA complex by NMR spectr
The three-dimensional structure of the vnd/NK-2 homeodomain-DNA complex by NMR spectroscopy.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--linkinghub.elsevier.com-ihub-images-PubMedLink.gif Related Articles The three-dimensional structure of the vnd/NK-2 homeodomain-DNA complex by NMR spectroscopy.
J Mol Biol. 1999 Jun 11;289(3):529-45
Authors: Gruschus JM, Tsao DH, Wang LH, Nirenberg M, Ferretti JA
The three-dimensional solution structure obtained by NMR of the complex formed between the uniformly singly15N and doubly13C/15N-labeled...
Analysis of SNARE complex/Synaptotagmin-1 Interactions by One-dimensional NMR Spectroscopy.
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