Table 2.
SAXS data collection parameters
| (A) Sample details | |||||
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| mtSSB (tetramer) | NEIL1 | mtSSB-DNA complex | NEIL1-mtSSB complex | NEIL1-DNA-mtSSB complex | |
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| Organism | Homo sapiens | Homo sapiens | Homo sapiens | Homo sapiens | Homo sapiens |
| Source | E. coli | E. coli | E. coli | E. coli | E. coli |
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| UniProt sequence ID (residues in constructs) | Q04837 (17–148) | Q96FI4 (1–390) | |||
| Extinction coefficients [A280, 0.1%(w/v)] | 1.313 | 0.725 | |||
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| SEC–SAXS column, 10/300 mm Superdex S200 | |||||
| Loading concentration (mg ml−1) | 7 | 11 | 8 | 10 | 10 |
| Injection volume (µl) | 300 | 300 | 300 | 300 | 300 |
| Flow rate (ml min−1) | 0.75 | 0.75 | 0.75 | 0.75 | 0.75 |
| Solvent (blanks taken from SEC flow through prior to elution of protein) | 25 mM HEPES pH 7.4, 5% glycerol, 300 mM NaCl, and 1 mM DTT | ||||
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| (B) Data collection parameters | |||||
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| Beamline | APS | APS | APS | APS | APS |
| Wavelength (Å) | 1.03 | 1.03 | 1.03 | 1.03 | 1.03 |
| Q Range (Å−1) | 0.0059–0.3892 | 0.0059–0.3898 | 0.0058–0.3587 | 0.0058–0.3746 | 0.0052–0.3606 |
| Temperature (°C) | 25 | 25 | 25 | 25 | 25 |
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| (C) Softwares used for data reduction, analysis and interpretation | |||||
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| SAXS data reduction | PRIMUS (ATSAS 2.8.1) | ||||
| Extinction coefficient estimate | ProtParam (Expasy) [83] | ||||
| Basic analyses: Guinier, P(r), MW (Vp, Vc) | PRIMUS (ATSAS 2.8.1), Scatter [43], BioXTAS RAW [47] | ||||
| Shape/bead modelling | DAMMIF and DAMAVER (ATSAS 2.8.1) | ||||
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| (C) Structural parameters | |||||
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| I(0) from P(r) | 44.88 | 51.49 | 22.74 | 102.8 | 116 |
| Rg (Å) from P(r) | 27.52 | 37.15 | 28.18 | 28.12 | 46.86 |
| I(0) from Guinier | 44.83±0.10 | 49.20±0.24 | 22.77±0.03 | 104.00±0.27 | 114.52±0.46 |
| Rg from Guinier | 27.44±1.21 | 33.04±2.46 | 28.40±0.86 | 28.48±0.80 | 44.83±1.80 |
| Dmax (Å) | 103.21 | 149.96 | 98.74 | 94.32 | 165.72 |
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| (D) Molecular weight determination (kDa) | |||||
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| Expected Theoretical (Expasy) | 60.78 | 44.72 | 78 | 105.5 | 123 |
| MW(Vp) | 61 | 46 | 79 | 60.3 | 146.6 |
| MW(Vc) | 59 | 36 | 71 | 55.4 | 134.3 |
| MW (MALS) | 59.8±1.67% | 48.5±2.23% | 69±1.46% | 58±1.54% | 129.6±1.54% |
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| (E) Modeling parameters | |||||
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| Symmetry | P4 | P1 | P4 | P1 | P1 |
| Particle anisometry | Oblate | Unknown | Oblate | Oblate, Prolate, Unknown | Oblate, Prolate, Unknown |
| # of modeling iterations | 10 | 10 | 10 | 10 | 10 |
| X2 of the model | 1.072 | 1.041 | 0.945 | 1.311(O) | 1.072(O) |
| 1.311(P) | 1.066(P) | ||||
| 1.312(U) | 1.069(U) | ||||