Probing slow dynamics in high molecular weight proteins by methyl-TROSY NMR spectroscopy: Application to a 723-residue enzyme

被引:179
作者
Korzhnev, DM
Kloiber, K
Kanelis, V
Tugarinov, V
Kay, LE
机构
[1] Univ Toronto, Protein Engn Network Ctr Excellence, Toronto, ON M5S 1A8, Canada
[2] Univ Toronto, Dept Med Genet, Toronto, ON M5S 1A8, Canada
[3] Univ Toronto, Dept Biochem, Toronto, ON M5S 1A8, Canada
[4] Univ Toronto, Dept Chem, Toronto, ON M5S 1A8, Canada
关键词
D O I
10.1021/ja039587i
中图分类号
O6 [化学];
学科分类号
0703 ;
摘要
A new CPMG-based multiple quantum relaxation dispersion experiment is presented for measuring millisecond dynamic processes at side-chain methyl positions in high molecular weight proteins. The experiment benefits from a methyl-TROSY effect in which cancellation of intramethyl dipole fields occurs, leading to methyl C-13-H-1 correlation spectra of high sensitivity and resolution (Tugarinov, V.; Hwang, P. M.; Ollerenshaw, J. E.; Kay, L. E. J. Am. Chem. Soc. 2003, 125, 10420-10428). The utility of the methodology is illustrated with an application to a highly deuterated, methyl-protonated sample of malate synthase G, an 82 kDa enzyme consisting of a single polypeptide chain. A comparison of the sensitivity obtained using the present approach relative to existing HSQC-type C-13 single quantum dispersion experiments shows a gain of a factor of 5.4 on average, significantly increasing the range of applications for this methodology.
引用
收藏
页码:3964 / 3973
页数:10
相关论文
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