Related ArticlesA simple method using 31P-NMR spectroscopy for the study of protein phosphorylation.
Brain Res Brain Res Protoc. 2000 Apr;5(2):182-9
Authors: Hirai H, Yoshioka K, Yamada K
Nonradioactive 31P-NMR spectroscopy has previously been used for the study of protein phosphorylations. However, the procedures does not seem to be easy for non-experts of this field, hence, this approach has not been widely used. We introduce here a simple protocol with 31P-NMR spectroscopy to study in vitro phosphorylation in receptor proteins. The effectiveness of this method was verified using synthetic peptides and recombinant proteins of the C-terminus of the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptor, whose phosphorylations are considered to have important roles in synaptic plasticity. We show that a decrease in the pH of the sample solution after the phosphorylation reaction is critical for the separation of the phosphorylation signals. In the analysis of the C-terminal portion of the GluR2 AMPA receptor, the phosphorylation sites of which had not hitherto been well clarified, we found the presence of at least three protein kinase C (PKC) phosphorylation sites. Furthermore, this method allows prediction of the origins of each of the phosphorylation peaks. Thus, the techniques we described here is useful for examination of protein phosphorylation and permits us to safely conduct repetitive experiments.
An NMR method to study protein-protein interactions.
An NMR method to study protein-protein interactions.
An NMR method to study protein-protein interactions.
Methods Mol Biol. 2012;757:129-37
Authors: Nishida N, Shimada I
Abstract
Specific interactions between proteins are a fundamental process underlying the various biological events, such as cell-cell contacts, signal transduction, and gene expression. Therefore, the structural investigations of protein-protein interactions provide useful information for understanding these events. We describe an NMR method, termed the cross-saturation...
nmrlearner
Journal club
0
09-13-2011 08:27 PM
A simple biosynthetic method for stereospecific resonance assignment of prochiral methyl groups in proteins
A simple biosynthetic method for stereospecific resonance assignment of prochiral methyl groups in proteins
Abstract A new method for stereospecific assignment of prochiral methyl groups in proteins is presented in which protein samples are produced using U-glucose and subsaturating amounts of 2-methyl-acetolactate. The resulting non-uniform labeling pattern allows proR and proS methyl groups to be easily distinguished by their different phases in a constant-time two-dimensional 1H-13C correlation spectra. Protein samples are conveniently prepared using the same media composition as the...
nmrlearner
Journal club
0
02-06-2011 07:42 PM
[NMR paper] High-throughput construction method for expression vector of peptides for NMR study s
High-throughput construction method for expression vector of peptides for NMR study suited for isotopic labeling.
Related Articles High-throughput construction method for expression vector of peptides for NMR study suited for isotopic labeling.
Protein Eng Des Sel. 2004 Apr;17(4):305-14
Authors: Tenno T, Goda N, Tateishi Y, Tochio H, Mishima M, Hayashi H, Shirakawa M, Hiroaki H
Fusion protein constructs for labeled peptides were generated with the 114 amino acid thioredoxin (TRX), coupled with the incorporation of a histidine tag for affinity...
nmrlearner
Journal club
0
11-24-2010 09:51 PM
[NMR paper] A simple protocol to study blue copper proteins by NMR.
A simple protocol to study blue copper proteins by NMR.
Related Articles A simple protocol to study blue copper proteins by NMR.
Eur J Biochem. 2003 Feb;270(4):600-9
Authors: Gelis I, Katsaros N, Luchinat C, Piccioli M, Poggi L
In the case of oxidized plastocyanin from Synechocystis sp. PCC6803, an NMR approach based on classical two and three dimensional experiments for sequential assignment leaves unobserved 14 out of 98 amino acids. A protocol which simply makes use of tailored versions of 2D HSQC and 3D CBCA(CO)NH and CBCANH leads to the...
nmrlearner
Journal club
0
11-24-2010 09:01 PM
[NMR paper] Insights into tyrosine phosphorylation control of protein-protein association from th
Insights into tyrosine phosphorylation control of protein-protein association from the NMR structure of a band 3 peptide inhibitor bound to glyceraldehyde-3-phosphate dehydrogenase.
Related Articles Insights into tyrosine phosphorylation control of protein-protein association from the NMR structure of a band 3 peptide inhibitor bound to glyceraldehyde-3-phosphate dehydrogenase.
Biochemistry. 1998 Jan 20;37(3):867-77
Authors: Eisenmesser EZ, Post CB
A protein-protein association regulated by phosphorylation of tyrosine is examined by NMR...
nmrlearner
Journal club
0
11-17-2010 11:06 PM
[NMR paper] The chemical shift index: a fast and simple method for the assignment of protein seco
The chemical shift index: a fast and simple method for the assignment of protein secondary structure through NMR spectroscopy.
Related Articles The chemical shift index: a fast and simple method for the assignment of protein secondary structure through NMR spectroscopy.
Biochemistry. 1992 Feb 18;31(6):1647-51
Authors: Wishart DS, Sykes BD, Richards FM
Previous studies by Wishart et al. have demonstrated that 1H NMR chemical shifts are strongly dependent on the character and nature of protein secondary structure. In particular, it has been...
nmrlearner
Journal club
0
08-21-2010 11:41 PM
A simple method for measuring signs of 1HN chemical shift differences between ground
Abstract NMR relaxation dispersion spectroscopy is a powerful method for studying protein conformational dynamics whereby visible, ground and invisible, excited conformers interconvert on the millisecond time-scale. In addition to providing kinetics and thermodynamics parameters of the exchange process, the CPMG dispersion experiment also allows extraction of the absolute values of the chemical shift differences between interconverting states,
ê
ê \Updelta ~
w
ê
ê , opening the way for structure determination of excited state conformers. Central to the goal of structural...
nmrlearner
Journal club
0
08-14-2010 04:19 AM
A simple method for amino acid selective isotope labeling of recombinant proteins in E. coli
A simple method for amino acid selective isotope labeling of recombinant proteins in E. coli
Kit I. Tong, Masayuki Yamamoto and Toshiyuki Tanaka
Journal of Biomolecular NMR; 2008; 42(1); pp 59-67
Abstract:
A simple and user-friendly method of labeling protein selectively with amino acids in vivo is introduced. This technique does not require the use of transaminase-deficient or auxotrophic strains. By manipulating the product feedback inhibitory loops of the E. coli amino acid metabolic pathways and, if necessary, by using enzyme inhibitors, proteins were labeled efficiently in vivo...
A simple method using 31P-NMR spectroscopy for the study of protein phosphorylation.
Linear Mode
