Table 1. Summary of sedimentation equilibrium data.
| Protein | Mr | ν (ml/g) | ρ (g/ml) | Mw | Fit variance | C range (μM) | Ka (M−1)a |
|---|---|---|---|---|---|---|---|
| Rna15b | 33 856 | 0.725 | 1.005 | 36 300 ± 2500 | 1.4 × 10−5 | 8–17 | — |
| Rna15NtDc | 11 476 | 0.721 | 1.005 | 11400 ± 400 | 2 × 10−5 | 25–140 | — |
| Rna15CtDd | 19 974 | 0.737 | 1.005 | 22 900 ± 200 | 2 × 10−5 | 18–100 | — |
| Rna14e | 83 025 | 0.730 | 1.007f | 145 000 ± 3000 | 2.6 × 10−5 | 0.66–2.52 | 2 × 106 ± 20% |
| Rna14–Rna15 (1:1)g | 114 058 | 0.729 | 1.005 | 140 000–205 000 | 3–5 × 10−5 | 1–11 | 6 × 105 ± 20% |
aThe equilibrium constants determined directly from absorbance data are expressed in the form of absorbance units termed Ka,abs. Actual equilibrium constants, expressed on a molar basis, Ka were converted from Ka,abs by the application of the expression Ka = ɛλlKa,abs/2 taking into account the extinction coefficient ελ of the sample and the optical pathlength of the cell (12 mm).
bRna15 molecular weight data were derived from global fitting of sedimentation equilibrium absorbance data (λ = 280 nm). The data were collected at three sample concentrations using rotor speeds of 14 000, 17 000 and 19 000. The final global fit incorporates data from nine independent measurements.
cRna15NtD molecular weight data were derived by global fitting of sedimentation equilibrium absorbance data (λ = 280 and 250 nm). The data were measured at nine sample concentrations using rotor speeds of 22 000, 26 000 and 30 000. The final global fit incorporates data from 15 independent measurements.
dRna15CtD molecular weight data were derived by global fitting of sedimentation equilibrium absorbance data (λ = 280 and 250 nm). The data were measured at nine sample concentrations using rotor speeds of 18 000, 20 000 and 22 000. The final global fit incorporates data from 15 independent measurements.
eRna14 molecular weight and equilibrium constant data were derived from global fitting of sedimentation equilibrium absorbance data measured at λ = 280 nm. The data were recorded at nine sample concentrations using rotor speeds of 8000, 10 000 and 12 000. The final global fit incorporates data from 12 independent measurements.
fAll sedimentation equilibrium experiments were carried out in 20 mM Tris–HCl, pH 7.8, 150 mM NaCl, 0.5 mM TCEP at 20°C except Rna14 where the NaCl concentration was 200 mM.
gRna14–Rna15 molecular weight and equilibrium constant data were derived from global fitting of sedimentation equilibrium absorbance data measured at λ = 280 and 250 nm. The data were recorded at nine sample concentrations using rotor speeds of 6000, 8000, 10 000 and 12 000. Data were fitted individually to determine the degree of concentration dependence and then separate global fits applied to data from each wavelength in order to derive equilibrium constants. The final value for Ka incorporates data from 27 independent measurements.