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So far high-resolution structure determination by nuclear magnetic resonance (NMR) spectroscopy has been limited to proteins <30 kDa, although global fold determination is possible for substantially larger proteins.
Rapid protein global fold determination using ultrasparse sampling, high-dynamic range artifact suppression, and time-shared NOESY.
RDC-ANALYTIC is a suite of programs for high-quality protein backbone fold determination from residual dipolar couplings (RDCs) (only two RDCs per residue are required) in one alignment medium, and a sparse set of nuclear Overhauser effect (NOE) data.
We illustrate this approach to protein fold determination on (15 N-labeled Eschericia coli acyl carrier protein using a limited set of (15 N-labeledd (1)H-(1)H dipolar couplings.
In structural studies of large proteins by NMR, global fold determination plays an increasingly important role in providing a first look at a target's topology and reducing assignment ambiguity in NOESY spectra of fully protonated samples.
Therefore, the inconsistent reduction in signal intensity may not be overcome in our experiments by pair-wise fold determination.
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The sensitivities increased from 57 and 59% in the case of T1 and Dukes' A, respectively, to 78% in the case of T4 and 90% in the case of Dukes' D. The intra-assay variance was evaluated by 18-fold determination of five samples (5 66 U ml−1), giving an average coefficient of variance (CV) of 7.9% (3.5 13.6%).
The 2-fold determination of ARI scores on two different days showed no differences at all, and the applied Dahlberg formula generated an error of zero [ 34].> -wrap-foot> ARI scores 0, no adhesive remains on tooth; 1, less than 50% of adhesive remains on tooth; 2, more than 50% of adhesive remains on tooth; 3, all adhesive remains on tooth.
All values were performed as 8-fold determinations.
Values are given as mean values ± standard deviation of 6-fold determinations.
Values for the respective protein compared to the vehicle-treated control ± standard deviation are depicted and represent 6-fold determinations.
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