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. Author manuscript; available in PMC: 2017 Feb 18.
Published in final edited form as: Nature. 2016 Aug 18;536(7616):304–308. doi: 10.1038/nature19071

Extended Data Table 2. Summary of Kinetic Constants For Multiple Turnover Experiments with Purified E2Ubc9-SUMOD68R Thioester.

(top) Catalytic constants for multiple turnover SUMO conjugation of PCNA to K127 at 30° C with 100 nM–5 µM purified E2Ubc9-SUMOD68R-Alexa488 thioester with and without a 1.5-fold excess of an additional molecule of SUMO, 1 nM E3 and 32 µM PCNA. Data show mean ± s.d. (n=3 technical replicates). (bottom) Catalytic constants for multiple turnover SUMO conjugation of PCNA to K164 at 30° C with 100 nM–5 µM purified E2Ubc9-SUMOD68R-Alexa488 thioester with and without a 1.5-fold excess of an additional molecule of SUMO, 1 nM E3 and 32 µM PCNA. Data show mean ± s.d. (n=3 technical replicates).

Sizl Isoform Additional SUMO Isoform Km (µM) kcat (s−1) kcat/ Km (M−1/s−1)
167–465 - 1.07 ± 0.10 1.27 ± 0.05 1.18 × 106 ± 0.12 × 106
167–449 - 1.23 ± 0.19 0.88 ± 0.05 7.15 × 105 ± 0.12 × 105
167–449 Δ18Smt3ΔGGHIS 1.84 ± 0.31 0.86 ± 0.06 4.68 × 105 ± 0.09 × 105
167–449 Δ18Smt3D68RΔGGHIS 1.08 ± 0.12 0.81 ± 0.03 7.49 × 105 ± 0.09 × 105
167–449-Δ18Smt3ΔGGHIS - 0.48 ± 0.15 0.64 ± 0.06 1.32 × 106 ± 0.43 × 106
167–449-Δ18Smt3D68RΔGGHIS - 4.40 ± 1.44 0.93 ± 0.18 2.11 × 105 ± 0.08 × 105
167–508 - 2.34 ± 0.58 0.54 ± 0.06 2.31 × 105 ± 0.06 × 105
167–508 Δ18Smt3ΔGGHIS 1.15 ± 0.14 0.89 ± 0.04 7.71 × 105 ± 0.10 × 105
167–508 Δ18Smt3D68RΔGGHIS 1.46 ± 0.18 0.75 ± 0.04 5.13 × 105 ± 0.07 × 105
167–508-Δ18Smt3ΔGGHIS - 0.29 ± 0.07 0.71 ± 0.04 2.43 × 106 ± 0.57 × 106
167–508-Δ18Smt3D68RΔGGHIS - 2.07 ± 0.29 0.70 ± 0.04 3.38 × 105 ± 0.05 × 105
Sizl Isoform Additional SUMO Isoform Km (µM) kcat (s−1) kcat/ Km (M−1/s−1)
167–465 - 0.70 ± 0.07 4.26 ± 0.14 6.09 × 106 ± 0.69 × 106
167–449 - 0.91 ± 0.12 2.52 ± 0.12 2.76 × 106 ± 0.39 × 106
167–449 Δ18Smt3ΔGGHIS 1.44 ± 0.21 2.47 ± 0.15 1.71 × 106 ± 0.27 × 106
167–449 Δ18Smt3D68RΔGGHIS 0.90 ± 0.07 2.45 ± 0.07 2.71 × 106 ± 0.22 × 106
167–449-Δ18Smt3ΔGGHIS - 0.23 ± 0.05 1.47 ± 0.08 6.48 × 106 ± 1.58 × 106
167–449-Δ18Smt3D68RΔGGHIS - 1.02 ± 0.19 1.29 ± 0.09 1.26 × 106 ± 0.25 × 106
167–508 - 0.91 ± 0.12 1.12 ± 0.05 1.29 × 106 ± 0.18 × 106
167–508 Δ18Smt3ΔGGHIS 0.62 ± 0.07 2.01 ± 0.07 3.22 × 106 ± 0.39 × 106
167–508 Δ18Smt3D68RΔGGHIS 0.65 ± 0.10 1.47 ± 0.08 2.27 × 106 ± 0.36 × 106
167–508-Δ18Smt3ΔGGHIS - 0.14 ± 0.03 1.77 ± 0.08 13.09 × 106 ± 2.81 × 106
167–508-Δ18Smt3D68RΔGGHIS - 0.76 ± 0.10 1.34 ± 0.06 1.77 × 106 ± 0.24 × 106