Datasheet for Conversion of CO2 into organic acids by engineered autotrophic yeast Baumschabl, M., Ata, Ö., Mitic, B.M., Lutz, L., Gassler, T., Troyer, C., Hann, S. and Mattanovich, D., 2022. Proceedings of the National Academy of Sciences, 119(47), p.e2211827119. Doi: https://doi.org/10.1073/pnas.2211827119 Table 1. Growth, itaconic and lactic acid production (Figure 1E-F) Growth (OD) Time 0 24 96 120 144 168 192 control 5.00 5.15 9.00 10.50 11.25 11.45 11.80 ita 4.67 4.03 7.87 9.50 10.17 11.23 11.13 lac 3.03 3.27 5.27 6.30 6.83 7.13 7.03 Production (mg/L) Time 0 24 96 120 144 168 192 control 0.00 0.00 0.00 0.00 0.00 0.00 0.00 ita 0.00 6.37 129.26 228.35 345.31 454.60 530.49 lac 0.00 0.00 70.22 127.47 146.95 164.05 195.93 Table 2. Growth and itaconic acid production (Figure 2). Growth (OD) Time 0 25 96 120.5 142.5 168 190 ita 4.00 4.40 8.75 9.75 11.05 13.70 14.70 ita mttA strong 4.13 3.73 4.63 4.80 5.07 5.07 5.10 ita mttA medium 4.07 3.97 7.89 8.43 9.17 10.53 11.45 ita mttA weak 4.43 4.23 8.17 9.37 10.23 11.87 13.07 control 4.30 4.90 8.60 9.70 10.60 12.60 13.80 Production [mg/L] itaconic acid Time 0 96 120.5 142.5 168 190 ita 0.00 124.72 205.14 302.29 412.07 525.14 ita mttA strong 0.00 72.66 101.10 126.43 164.60 194.53 ita mttA medium 0.00 214.76 341.00 471.39 634.47 754.81 ita mttA weak 0.00 132.26 229.68 327.75 429.67 562.18 Table 3. Growth and lactic acid consumption (Figure 3). Growth (OD) Time 0 25 77.5 122 174 ldh 3.26 4.79 7.21 7.83 7.55 ldh dCYB2 3.30 3.46 3.87 3.91 4.11 dCYB2 3.49 3.11 2.96 2.87 2.85 Lactic acid consumption [g/L] Time 0 25 77.5 122 174 ldh 9.11 7.72 5.44 4.63 4.18 ldh dCYB2 9.11 8.72 8.35 8.13 7.76 dCYB2 9.11 9.05 9.14 9.19 9.10 Table 4. Growth, lactic and glycolic acid production (Figure 4). Growth (OD) Time 0.00 20.50 93.50 141.50 165.00 189.00 256.50 ldh 3.03 3.27 5.27 6.30 6.83 7.13 7.03 ldh dCYB2 3.20 3.25 5.50 6.90 7.58 8.00 9.23 dCYB2 3.23 3.13 5.83 7.43 8.43 9.03 10.50 Production [mg/L] Lactic acid Time 0.00 20.50 93.50 141.50 165.00 189.00 256.50 ldh 0.00 0.00 70.22 127.47 146.95 164.05 195.93 ldh dCYB2 0.00 0.00 126.23 256.68 292.13 325.46 356.29 dCYB2 0.00 0.00 0.00 0.00 0.00 0.00 0.00 Production [mg/L] Glycolic acid Time 0 20.5 93.5 141.5 165 189 256.5 ldh 0 0 0 0 0 0 0 ldh dCYB2 0 0 39.1793422 69.2718148 83.1941962 100.076249 148.963004 dCYB2 0 0 44.8004804 84.3012772 106.017956 132.449979 197.685748 Table 5. Labeling degree of the produced organic acid (Figure 5). Labeling Deg (%) Time [day] 3 5 8 ita mttA medium 35.40 23.33 18.49 ΔRuBisCO control ita 79.67 75.18 ldh dCYB2 13.48 10.77 10.11 ΔRuBisCO control lac 78.20 67.57 Table 6. Growth, itaconic and lactic acid production (Figure 6). Growth (OD) 5 % CO2 Time 0.00 24 48 96 120 144 168 192 ita mttA medium OD4 3.85 3.80 4.50 6.00 6.45 7.10 7.50 8.05 ita mttA medium OD10 11.13 10.25 10.88 12.75 13.00 14.25 15.63 14.00 ita mttA medium OD20 20.00 17.25 18.50 20.00 20.50 21.50 23.25 23.38 Growth (OD) 10% CO2 Time 0 24 48 72 96 120 144 168 192 ita mttA medium OD4 4.40 4.40 4.95 6.00 6.00 7.20 7.15 8.38 8.75 ita mttA medium OD10 11.88 10.50 13.75 11.75 12.25 13.50 14.13 14.75 14.00 ita mttA medium OD20 21.63 20.63 22.25 19.38 19.38 20.13 21.63 21.38 21.50 Growth (OD) 5% CO2 Time 0 22 44 70 92 116 139 164 189 ldh dCYB2 OD4 4.30 4.95 6.40 6.22 7.25 6.78 7.44 8.10 8.29 ldh dCYB2 OD8 8.60 9.40 12.05 12.52 13.84 13.56 14.03 15.07 15.16 ldh dCYB2 OD12 12.25 12.85 17.75 17.38 19.88 18.38 18.63 19.63 19.50 ldh dCYB2 OD16 17.14 18.46 21.38 20.50 24.25 23.25 22.00 22.13 23.00 Growth lac 10% CO2 Time 0 21 45.5 71 94 121.5 142.5 166 191.5 ldh dCYB2 OD4 4.65 5.65 7.25 8.10 7.82 9.32 9.79 10.36 10.92 ldh dCYB2 OD8 8.15 9.50 12.05 13.94 13.56 16.20 16.57 17.14 16.39 ldh dCYB2 OD12 13.09 16.10 17.16 18.66 19.65 22.26 21.52 22.26 20.52 ldh dCYB2 OD16 17.04 20.25 21.02 22.51 22.89 25.99 25.37 26.12 24.13 Production [mg/L] itaconic acid 5% CO2 Time 0.00 24 48 96 120 144 168 192 ita mttA medium OD4 0.00 4.97 20.07 104.51 186.53 286.29 417.96 553.38 ita mttA medium OD10 0.00 10.46 41.16 216.78 348.71 521.09 704.30 908.68 ita mttA medium OD20 0.00 11.42 55.08 250.31 385.86 482.31 782.99 1025.66 Production [mg/L] itaconic acid 10% CO2 Time 0 24 48 72 96 120 144 168 192 ita mttA medium OD4 0 11.51 52.42 134.32 250.55 423.76 535.94 705.85 886.39 ita mttA medium OD10 0 33.44 120.65 301.66 535.17 852.21 1064.25 1349.70 1611.05 ita mttA medium OD20 0 59.12 202.88 411.03 689.18 1086.69 1353.11 1707.48 1979.99 Production [mg/L] lactic acid 5% CO2 Time 0 22 44 70 92 116 139 164 189 ldh dCYB2 OD4 0 0.00 25.52 79.79 181.78 330.17 462.69 579.77 640.94 ldh dCYB2 OD8 0 14.92 44.72 124.31 260.68 416.45 446.83 389.99 316.38 ldh dCYB2 OD12 0 19.60 56.95 142.31 253.30 272.53 249.03 213.54 182.71 ldh dCYB2 OD16 0 22.43 67.10 162.35 258.67 232.86 217.47 194.86 172.94 Production [mg/L] lactic acid 10% CO2 Time 0 21 45.5 71 94 121.5 142.5 166 191.5 ldh dCYB2 OD4 0 0 20.48 66.30 132.66 226.66 297.36 361.14 413.11 ldh dCYB2 OD8 0 0 36.39 93.33 174.70 283.77 346.56 358.73 330.51 ldh dCYB2 OD12 0 15.09 50.21 111.44 192.28 290.09 307.19 283.59 260.00 ldh dCYB2 OD16 0 17.97 58.69 121.65 198.53 279.36 279.26 252.93 239.13 Production [mg/L] glycolic acid 5% CO2 Time 0 22 44 70 92 116 139 164 189 ldh dCYB2 OD4 0 0 18.01 34.86 51.00 66.07 81.12 97.96 116.78 ldh dCYB2 OD8 0 0 28.38 53.96 78.64 99.69 123.00 143.44 176.90 ldh dCYB2 OD12 0 0 32.26 59.80 84.17 104.09 122.99 146.55 168.90 ldh dCYB2 OD16 0 0 36.49 68.26 96.01 117.13 134.11 146.14 164.64 Production [mg/L] glycolic acid 10% CO2 Time 0 21 45.5 71 94 121.5 142.5 166 191.5 ldh dCYB2 OD4 0 0 0 19.03 27.27 39.54 48.30 59.05 72.19 ldh dCYB2 OD8 0 0 0 26.23 39.06 56.94 71.41 86.50 96.37 ldh dCYB2 OD12 0 0 17.02 31.34 46.64 66.75 80.38 93.46 112.71 ldh dCYB2 OD16 0 0 19.25 35.33 50.73 71.84 84.69 95.71 111.24 Table 7. Growth, itaconic and lactic acid production in the bioreactor cultivations (Figure 7) Growth (OD) Time 0 23.68 28.53 48.23 52.08 70.93 74.53 120.53 124.67 144.1 148.63 173.63 192.69 DO20% 34.4 32.2 28.6 31.4 29.6 29.6 29 29.8 28 31.2 28.6 28.6 28.4 Time 0 18.5 22 45 47 91 94 118 137 141 166 184 DO 8% 21.6 20.7 19.7 20 19.9 20.1 19.5 18.9 21 18.6 19.2 18.2 Production [mg/L] itaconic acid Time 0 23.68 28.53 48.23 52.08 70.93 74.53 120.53 124.67 144.1 148.63 173.63 192.69 DO20% 0 75.56 99.20 115.86 116.63 129.25 130.83 163.98 171.34 194.10 202.43 219.60 241.67 Time 0 18.5 22 45 47 91 94 118 137 141 166 184 DO 8% 0 53.57 67.46 158.50 150.43 293.37 303.56 372.94 424.46 432.33 490.29 527.72 Growth (OD) Time 0 18.33 22 41 65.33 120.45 161.40 189.00 215.50 234.60 304.80 328.30 352.30 376.80 405.33 473.25 497.17 21% Oxygen 2.28 2.22 2.36 2.60 3.15 4.02 4.21 4.34 4.63 4.69 4.79 5.07 4.98 5.04 5.17 5.33 5.36 5% Oxygen 2.16 2.1 2.31 2.46 3.25 4.28 5.11 5.36 6.13 6.38 8.03 9.02 9.54 9.98 11.34 12.22 12.48 Production [mg/L] lactic acid 21% Oxygen 0 0 0 8.29 14.96 44.74 69.29 85.62 100.50 109.04 153.28 167.34 181.66 196.13 213.57 253.80 266.21 5% Oxygen 0 0 0 9.18 14.67 23.25 31.95 37.72 44.91 48.82 72.42 79.05 88.38 95.44 104.33 133.56 139.99 Production [mg/L] glycolic acid 21% Oxygen 0 0 0 8.98 13.84 28.41 39.91 48.41 57.04 62.87 86.78 94.87 102.48 110.89 120.28 144.56 151.34 5% Oxygen 0 0 0 5.08 5.68 7.61 11.07 13.00 14.89 15.78 22.24 23.72 25.74 28.10 30.38 38.60 40.31