TABLE 2.

Summary of NMR of Hsc70-PP5

Domain	Molecular mass	τc at 30 °C Ca	1HN HSQC LWb	1HN TROSY LWb	15N TROSY LWb	TROSY transfer efficiencyc	Rel. peak heightd
	kDa	ns	Hz	Hz	Hz	%	a.u.
Tail	10	7	20	16	0.5	39	1.0 × 10
SBD	25	14	40	32	0.9	15	1.9 × 10−1
NBD	45	23	65	51	1.5	4.7	3.8 × 10−2
Hsc70	70	40	112	90	2.5	0.5	2.3 × 10−3
PP5+tail	70	40	112	90	2.5	0.5	2.3 × 10−3
PP5+SBD	85	45	126	100	2.8	0.3	1.1 × 10−3
PP5+NBD	105	55	154	122	3.5	0.07	2.4 × 10−4
PP5+Hsc70	130	70	196	156	4.4	0.01	2.6 × 10−5

^a The rotational correlation time τ_c was estimated from the molecular mass following Ref. 38.

^b The average amide proton and nitrogen line widths (LW) at 800 MHz were calculated from coordinates of the crystal structure of ubiquitin as a model, using different rotational correlation times. The calculations took into account all dipole-dipole interactions with all magnetic nuclei in the molecule, and ¹H CSA or ¹⁵N CSA relaxation. ¹H-¹⁵N dipolar/¹H CSA or ¹H-¹⁵N/¹⁵N CSA cross-correlated R₂ relaxation was taken into account for the columns marked “TROSY.” We assumed uniform ¹⁵N labeling, no ¹³C, or ²H labeling.

^c In TROSY, there are three transfer periods with ¹HN coherence, which all are tuned to 1/2J_NH (5 ms). The transfer efficiency is reduced by ¹H R₂ relaxation. In total, I = I₀ (exp(−3.1416 × LW × 0.005))³. The relevant line widths during these transfers are listed in the column marked “¹HN HSQC LW.” Rel., relative.

^d The peak height for 10 kDa was taken as a standard. The peak heights for other molecular weights were computed by taking the ratio of the relevant transfer efficiencies divided by the ratio of the relevant ¹H TROSY linewidths (the latter affects peak height during data acquisition). The effect of increasing ¹⁵N line width on the TROSY peak intensity is minimal because of the short ¹⁵N acquisition time used and was not included in the calculation; a.u., absorbance units.