Amino acid-selective isotope labeling of proteins for nuclear magnetic resonance study: proteins secreted by Brevibacillus choshinensis.
Here we report the first application of amino acid-type selective (AATS) isotope labeling of a recombinant protein secreted by Brevibacillus choshinensis for a nuclear magnetic resonance (NMR) study. To prepare the 15N-AATS-labeled protein, the transformed B. choshinensis was cultured in 15N-labeled amino acid-containing C.H.L. medium, which is commonly used in the Escherichia coli expression system. The analyses of the 1H-15N heteronuclear single quantum coherence (HSQC) spectra of the secreted proteins with a 15N-labeled amino acid demonstrated that alanine, arginine, asparagine, cysteine, glutamine, histidine, lysine, methionine, and valine are suitable for selective labeling, although acidic and aromatic amino acids are not suitable. The 15N labeling for glycine, isoleucine, leucine, serine, and threonine resulted in scrambling to specific amino acids. These results indicate that the B. choshinensis expression system is an alternative tool for AATS labeling of recombinant proteins, especially secretory proteins, for NMR analyses.
Tanio
Michikazu
M
Mitsubishi Kagaku Institute of Life Sciences (MITILS), Molecular Structure Research Group, 11 Minamiooya, Machida, Tokyo 194-8511, Japan.
Tanaka
Rikou
R
Tanaka
Takeshi
T
Kohno
Toshiyuki
T
D000596:Amino Acids / Q000737:chemistry* / Q000378:metabolism; D006094:Gram-Positive Bacteria / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D007553:Isotope Labeling; D009587:Nitrogen Isotopes / Q000737:chemistry; D019906:Nuclear Magnetic Resonance, Biomolecular; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D022021:Tacrolimus Binding Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics
D000596:Amino Acids; D009587:Nitrogen Isotopes; D011994:Recombinant Proteins; D022021:Tacrolimus Binding Proteins