Table 1.

Analysis and deconvolution of S₀ and S₁ solid-state NMR spectra of membrane-reconstituted Fgp41

Fgp41 labeling	(ΔS/S0)exp (integrated)c	S0 spectral deconvolution a				Fraction of calculated S0 intensity b
		Peak shift (ppm) d		Peak width (ppm) e	Intensity (fraction of total)	Labeled	Nat. abund. Fgp41	Nat. abund. lipid	Labeled in N- and C- helices	(ΔS/S0)calc,f
1-13C, 15N Leu	0.12	181.3	helix	2.8	0.15
		178.5		2.8	0.60	0.86	0.07	0.07	0.68	0.15
		175.3	β	3.7	0.25
1-13C Phe + 15N Leu	0.15	183.2	helix	3.0	0.02
		177.1		5.0	0.51	0.42	0.31	0.27	0	0.24
		172.7	β	3.5	0.47
1-13C Val + 15N Phe	0.07	182.1	helix	2.7	0.02
		177.7		3.9	0.76	0.69	0.19	0.12	0.43	0.07
		173.1	β	3.3	0.22
1-13C Val + 15N Gly	0.08	178.6	helix	2.9	0.37
		177.0		2.0	0.20	0.64	0.17	0.19	0.40	0.07
		174.4	β	4.9	0.43
1-13C Ala + 15N Gly	0.08					0.79	0.12	0.09	0.34	0.05
1-13C Gly + 15N Leu	0.11					0.77	0.15	0.08	0.14	0.06

^aSpectral deconvolution was done with three Gaussian lineshapes whose peak shifts, linewidths, and intensities were independently varied until there was minimal difference between the sum of the lineshapes and the experimental lineshape. For all cases, there was excellent agreement between the best-fit deconvolution sum lineshape and experimental lineshape, see Supporting Information. Deconvolution was not meaningful for the 1-¹³C Ala and 1-¹³C Gly samples because the S₀ spectra were broad and relatively featureless and the deconvolutions were dominated by a lineshape with ~7 ppm linewidth.

^bSpectral intensities were calculated with the following considerations: (1) 100% labeling of the Fgp41 residues corresponding to the labeled amino acid(s) with no scrambling to other amino acid types; (2) 1.0 relative intensity for each labeled ¹³CO; (3) 0.011 relative intensity for each natural abundance ¹³CO; (4) the Fgp41 natural abundance signal was the sum from backbone ¹³COs and Asn, Asp, Gln, and Glu sidechain ¹³COs; and (5) the lipid natural abundance signal was calculated using the experimental Fgp41:total lipid mol ratios. The specific ratio in each sample: 1-¹³C, ¹⁵N Leu, 0.011; 1-¹³C Phe + ¹⁵N Leu, 0.012; 1-¹³C Val + ¹⁵N Phe, 0.016; 1-¹³C Val + ¹⁵N Gly, 0.009; 1-¹³C Ala + ¹⁵N Gly, 0.013; and 1-¹³C Gly + ¹⁵N Leu, 0.019. The labeled ¹³CO fraction in N- and C- helices was based on the red and green regions in Fig. 1a.

^cThe typical uncertainty of (ΔS/S₀)^exp was ±0.02 as determined from the standard deviation of integrals of regions of the S₀ and S₁ spectra which contained noise rather than signal.

^dThe reasons for assignment of peaks to specific conformations are provided in the main text.

^eFull-width at half-maximum linewidth.

^fThe (ΔS/S₀)^calc were based on: (1) the fraction of the S₀ signal from labeled ¹³COs directly bonded to labeled ¹⁵Ns; and (2) a S₁/S₀ intensity ratio for these ¹³COs of 0.70 (1 ms dephasing time) or 0.85 (2 ms dephasing time). These ratios were based on experimental REDOR data of crystalline amino acid samples as well as simulations. The 1 ms dephasing time was used for the Fgp41 sample labeled with 1-¹³C, ¹⁵N Leu and the 2 ms dephasing time was used for the other Fgp41 samples.