. Author manuscript; available in PMC: 2011 Sep 22.

Published in final edited form as: J Am Chem Soc. 2010 Sep 22;132(37):13026–13045. doi: 10.1021/ja105485b

Table 1.

SVD analysis of ¹D_NH RDCs measured for residues of the EIN domain in intact EI and the EI-HPr complex.^a

	Number of RDCs	Euler angles (°)^b			D_a^NH (Hz)	η	RDC R-factor^c
	Number of RDCs	ϕ	θ	ψ	D_a^NH (Hz)	η	RDC R-factor^c
A. Free EI
Fits to A state (free EIN conformation)^d
EIN^α/β	29	231	13	139	13.9	0.56	15.6
EIN^α	29	249 (230)	10 (8)	113 (136)	14.7	0.49	18.4
EIN global	58	242 (235)	10 (10)	122 (132)	14.6 (14.6)	0.49 (0.50)	18.0 (17.5)
Fits to B state (orientation in crystal structure of phosphorylated EI from E. coli)
EIN^α/β	29	231	13	139	13.9	0.56	15.6
EIN^α	29	279	33	149	14.7	0.49	18.4
EIN global	58	250	28	147	13.0	0.32	36.8
Fits to orientation in crystal structure of EI from Staph. carnosus
EIN^α/β	29	231	13	139	13.9	0.56	15.6
EIN^α	29	288	15	114	14.7	0.49	18.4
EIN global	58	243	11	141	14.4	0.31	25.5
B. EI-HPr complex
Fits to A state (free EIN conformation)^d
EIN^α/β	25	242	11	121	13.7	0.63	12.1
EIN^α	18	244 (216)	12 (8)	118 (144)	14.0	0.65	20.3
EIN global	43	243 (238)	10 (10)	118 (122)	13.9 (13.9)	0.63 (0.64)	16.2 (15.5)
Fits to B state (orientation in crystal structure of phosphorylated EI from E. coli)
EIN^α/β	25	242	11	121	13.7	0.63	12.1
EIN^α	18	275	32	146	14.0	0.65	20.3
EIN global	43	257	30	120	12.3	0.36	34.6
Fits to orientation in crystal structure of EI from Staph. carnosus
EIN^α/β	25	242	11	121	13.5	0.62	12.4
EIN^α	18	284	13	112	14.0	0.65	20.4
EIN global	43	242	12	132	13.3	0.40	23.7

The coordinates used for SVD analysis are those of EIN in the EIN-HPr complex (3EZA).¹³ The EIN^α and EIN^α/β subdomains are then fitted onto the X-ray coordinates of E. coli phosphorylated EI (2HWG)¹⁸ and Staph carnosus (2HRO)¹⁶ EI. Thus the comparisons reflect only the relative orientation of the EIN^α and EIN^α/β subdomains and are not influenced by variations in the atomic coordinates for the different structures. SVD analysis was carried out in Xplor-NIH.³⁵

For ease of comparison, the EIN^α/β subdomain is positioned in the same molecular frame throughout. The convention used for the Euler angles is the x convention of Goldstein et al.⁶⁴ ϕ describes the rotation about the z axis, θ the rotation about the resulting x axis, and ψ the rotation about the resulting z axis.

The RDC R-factor⁵⁸ is defined as R_inf = [<(D_obs-D_calc)²>/(2<D_obs²>)]^1/2 where D_obs and D_calc are the observed and calculated RDC values.

The values in parentheses refer to the values obtained using the NMR EIN^α and EIN^α/β coordinates in the relative orientation found in the crystal structure of free EIN (1ZYM).¹¹ (i.e. these hybrid coordinates were generated by best-fitting the EIN^α and EIN^α/β subdomains of the 3EZA NMR structure individually on to the corresponding subdomains of 1ZYM. The overall Cα rms difference for EIN^α + EIN^α/β between the 3EZA and 1ZYM orientations of the two subdomains is 0.7 Å, and the rotation and Cα rms displacement between the EIN^α subdomains when the coordinates are best-fitted to the EIN^α/β subdomains are 5.7° and 1.8 Å, respectively. These differences are within the experimental error of the current data.