NMR assignment methods for the aromatic ring resonances of phenylalanine and tyrosine residues in proteins.
The unambiguous assignment of the aromatic ring resonances in proteins has been severely hampered by the inherently poor sensitivities of the currently available methodologies developed for uniformly 13C/15N-labeled proteins. Especially, the small chemical shift differences between aromatic ring carbons and protons for phenylalanine residues in proteins have prevented the selective observation and unambiguous assignment of each signal. We have solved all of the difficulties due to the tightly coupled spin systems by preparing regio-/stereoselectively 13C/2H/15N-labeled phenylalanine (Phe) and tyrosine (Tyr) to avoid the presence of directly connected 13C-1H pairs in the aromatic rings. The superiority of the new labeling schemes for the assignment of aromatic ring signals is clearly demonstrated for a 17 kDa calcium binding protein, calmodulin.
Torizawa
Takuya
T
CREST-JST and Graduate School of Science, Tokyo Metropolitan University, 1-1 Minami-ohsawa, Hachioji, 192-0397, Japan.
Ono
Akira Mei
AM
Terauchi
Tsutomu
T
Kainosho
Masatsune
M
D002247:Carbon Isotopes; D009682:Magnetic Resonance Spectroscopy / Q000379:methods* / Q000592:standards; D015394:Molecular Structure; D010649:Phenylalanine / Q000737:chemistry*; D011506:Proteins / Q000737:chemistry*; D011522:Protons; D012015:Reference Standards; D014443:Tyrosine / Q000737:chemistry*
D002247:Carbon Isotopes; D011506:Proteins; D011522:Protons; D014443:Tyrosine; D010649:Phenylalanine