Related ArticlesAmino acid type selective isotope labelling of the multidrug ABC transporter LmrA for solid-state NMR studies.
FEBS Lett. 2004 Jun 18;568(1-3):117-21
Authors: Mason AJ, Siarheyeva A, Haase W, Lorch M, van Veen H, Glaubitz C
The ABC transporter LmrA in Lactococcus lactis confers resistance to a wide range of antibiotics and cytotoxic drugs and is a functional homologue of P-glycoprotein. Recently, solid-state NMR methods have shown potential for structural- and non-perturbing, site directed functional studies. These experiments require isotopic labelling of selected sites. We have developed a strategy to produce large quantities of selectively labelled LmrA reconstituted at a high density in lipid membranes. This makes the 64 kDa integral membrane protein LmrA and therefore the ABC transporter superfamily accessible to NMR analysis.
Amino acid selective unlabeling for sequence specific resonance assignments in proteins
Amino acid selective unlabeling for sequence specific resonance assignments in proteins
Abstract Sequence specific resonance assignment constitutes an important step towards high-resolution structure determination of proteins by NMR and is aided by selective identification and assignment of amino acid types. The traditional approach to selective labeling yields only the chemical shifts of the particular amino acid being selected and does not help in establishing a link between adjacent residues along the polypeptide chain, which is important for sequential assignments. An alternative...
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03-20-2012 12:42 AM
A simple protocol for amino acid type selective isotope labeling in insect cells with improved yields and high reproducibility
A simple protocol for amino acid type selective isotope labeling in insect cells with improved yields and high reproducibility
Abstract An easy to use and robust approach for amino acid type selective isotope labeling in insect cells is presented. It relies on inexpensive commercial media and can be implemented in laboratories without sophisticated infrastructure. In contrast to previous protocols, where either high protein amounts or high incorporation ratios were obtained, here we achieve both at the same time. By supplementing media with a well considered amount of yeast extract,...
Suppression of isotope scrambling in cell-free protein synthesis by broadband inhibition of PLP enymes for selective 15N-labelling and production of perdeuterated proteins in H2O
Suppression of isotope scrambling in cell-free protein synthesis by broadband inhibition of PLP enymes for selective 15N-labelling and production of perdeuterated proteins in H2O
Abstract Selectively isotope labelled protein samples can be prepared in vivo or in vitro from selectively labelled amino acids but, in many cases, metabolic conversions between different amino acids result in isotope scrambling. The best results are obtained by cell-free protein synthesis, where metabolic enzymes are generally less active, but isotope scrambling can never be suppressed completely. We show that...
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02-16-2011 09:34 PM
[NMR paper] Site-selective screening by NMR spectroscopy with labeled amino acid pairs.
Site-selective screening by NMR spectroscopy with labeled amino acid pairs.
Related Articles Site-selective screening by NMR spectroscopy with labeled amino acid pairs.
J Am Chem Soc. 2002 Mar 20;124(11):2446-7
Authors: Weigelt J, van Dongen M, Uppenberg J, Schultz J, Wikström M
A new method for site-selective screening by NMR is presented. The core of the new method is the dual amino acid sequence specific labeling technique. Amino acid X is labeled with (13)C and amino acid Y is labeled with (15)N. Provided only one XY pair occurs in the...
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11-24-2010 08:49 PM
[NMR paper] NMR investigation of the multidrug transporter EmrE, an integral membrane protein.
NMR investigation of the multidrug transporter EmrE, an integral membrane protein.
Related Articles NMR investigation of the multidrug transporter EmrE, an integral membrane protein.
Eur J Biochem. 1998 Jun 15;254(3):610-9
Authors: Schwaiger M, Lebendiker M, Yerushalmi H, Coles M, Gröger A, Schwarz C, Schuldiner S, Kessler H
EmrE is an Escherichia coli multidrug transport protein that confers resistance to a wide range of toxicants by active transport across the bacterial cell membrane. The highly hydrophobic polytopic integral membrane...
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11-17-2010 11:06 PM
[NMR paper] Cell-free synthesis and amino acid-selective stable isotope labeling of proteins for
Cell-free synthesis and amino acid-selective stable isotope labeling of proteins for NMR analysis.
Related Articles Cell-free synthesis and amino acid-selective stable isotope labeling of proteins for NMR analysis.
J Biomol NMR. 1995 Sep;6(2):129-34
Authors: Kigawa T, Muto Y, Yokoyama S
For the application of multidimensional NMR spectroscopy to larger proteins, it would be useful to perform selective labeling of one of the 20 amino acids. For some amino acids, however, amino acid metabolism drastically reduces the efficiency and selectivity...
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08-22-2010 03:50 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...
Amino acid type selective isotope labelling of the multidrug ABC transporter LmrA for
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