Related ArticlesSTD-NMR based protein engineering of the unique arylpropionate-racemase AMDase G74C.
Chembiochem. 2015 Jun 23;
Authors: Gassmeyer SK, Yoshikawa H, Enoki J, Hülsemann N, Stoll R, Miyamoto K, Kourist R
Abstract
Structure-guided protein engineering achieved a variant of the unique racemase AMDase G74C with 40-fold increased activity in the racemisation of several arylali¬phatic carboxylic acids. By saturation-transfer-difference (STD) NMR spectroscopy, substrate binding during catalysis was investigated. All atoms of the substrate showed interactions with the enzyme. STD-NMR measurements revealed distinct nuclear Overhauser effects in experiments with and without molecular conversion. The spectroscopic analysis led to the identification of several amino acid residues whose variation increased the activity of G74C. While single-amino acid exchanges increased the activity moderately, structure-guided saturation mutagenesis yielded a quadruple mutant with a 40 times higher reaction rate. This study presents STD-NMR as versatile tool for the analysis of enzyme-substrate interactions in catalytically competent systems and for the guidance of protein engineering.
PMID: 26109370 [PubMed - as supplied by publisher]
Protein engineering: The power of four - Nature.com
Protein engineering: The power of four - Nature.com
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Protein engineering: The power of four
Nature.com
The final structure of the dimer of dimers, which they called 'rocker', was characterized by X-ray crystallography, and a variety of NMR techniques, which, in combination with molecular dynamic simulations, indicated that the peptide assembly did ...
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http://pubs.acs.org/appl/literatum/publisher/achs/journals/content/jacsat/0/jacsat.ahead-of-print/ja408988z/aop/images/medium/ja-2013-08988z_0015.gif
Journal of the American Chemical Society
DOI: 10.1021/ja408988z...
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The calcium regulatory protein calmodulin (CaM) plays a role as an on-off switch in the activation of many enzymes and proteins. CaM has a dumbbell shaped structure with two folded domains, which are connected by a flexible linker in solution. The calmodulin-binding domains of the target proteins are contained in 20 residue long amino acid sequences, that...
STD-NMR based protein engineering of the unique arylpropionate-racemase AMDase G74C.
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