Table 3.
Structural statistics for refinement of EI and EI-HPr structures based on SAXS/WAXS and RDC data.a
| free EI (100) |
EI-HPr complex | ||
|---|---|---|---|
| Cluster 1 (33) |
Cluster 2 (67) |
||
| SAXS/WAXS χ2 b | |||
| q = 0.014/0.022→0.44 Å-1 | 0.30±0.05 | 0.63±0.11 | 0.76±0.07 |
| q complete range predicted | 2.25±0.62 | 0.45±0.06 | 0.48±0.06 |
| q complete range fitted | 1.34±0.27 | 0.45±0.07 | 0.48±0.06 |
| SANS χ2 c | 0.41±0.08 | 1.38±0.51 | 2.97±0.62 |
| RDC R-factor (%)d | 18.07±0.02 | 16.30±0.03 | 16.25±0.02 |
| RDC DaNH (Hz) | 14.5±0.1 | 13.73±0.05 | 13.83±0.08 |
| RDC η | 0.49±0.00 | 0.63±0.00 | 0.63±0.00 |
| Coordinate precisione | |||
| EIN Cα rms displacement (Å) | 1.3±0.7 | 2.2±1.2 | 2.2±1.3 |
| rotation (deg.) | 4.2±2.9 | 3.0±2.0 | 4.2±3.0 |
| translation (Å) | 3.5±2.3 | 3.0±1.8 | 3.9±2.8 |
The final number of simulated annealing structures is given in parentheses. 100 final simulated annealing structures were calculated for free EI and the EI-HPr complex. Using a cutoff of 3 Å, the EI-HPr structures cluster into two groups with 33 structures in cluster 1 and 67 in cluster 2.
In these structure calculations only the SAXS/WAXS data up to q = 0.44 Å-1 are used. The lowest experimental q values collected for the EI and EI-HPr data are 0.014 and 0.022 Å-1, respectively. Two χ2 values are reported for the complete SAXS/WAXS curve up to q = 0.8 Å-1 for free EI and 0.95 for the EI-HPr complex: The first set of values, denoted as ‘q complete range predicted’, are obtained by only fitting the data up to q = 0.44 Å-1; these χ2 values therefore report on how well the structures obtained using only data up to q = 0.44 Å-1 predict the data from q = 0.44 to 0.8 Å-1 for free EI and from q = 0.44 to 0.95 Å-1 for the EI-HPr complex. The second set of values, denoted as ‘q complete range fitted’, are obtained when the complete SAXS/WAXS data are used in the fitting procedure (q = 0.014→0.8 Å-1 for EI and 0.022→0.95 Å-1 for EI-HPr).
The SANS data were collected from q = 0.035→0.20 Å-1 for 2H-EI and from q = 0.028→0.21 Å-1 for 2H-EI/1H-HPr.
The target values for the RDC R-factors (i.e. the values obtained by SVD analysis using the NMR coordinates of EIN alone; cf Table 1) are 18.02 and 16.23%, respectively.
The precision with which the position of the EIN domain or the EIN domain plus HPr is determined relative to the dimeric EIC domain (in the context of intact EI) is defined by three parameters: the average atomic Cα rms displacement, and the average rotation and translation (along the axis joining the center of masses of the two EIN, or EIN plus HPr, domains being compared) relative to the EIN domain (or EIN plus HPr) of the restrained regularized mean coordinates. All structures are first best-fit to the EIC domain (residues 262-573) whose coordinates are fixed and identical in all cases.