Table 3.
Quantification of oligosaccharides produced by B. megaterium levansucrase variants from 0.5 m sucrose.
| Variant | Position | 1-kestose mm | 6-Kestose mm | 6-nystose mm | Unknown Disaccharidea | Neokestosea | ||
|---|---|---|---|---|---|---|---|---|
| 370 | 373 | 419 | ||||||
| WT | R | K | F | 1.68 ± 0.27 | 0.35 ± 0.01 | 1.29 ± 0.14 | 1.00 | 1.00 |
| 1H11 | H | K | F | 4.99 ± 0.30 | 0.54 ± 0.06 | 1.10 ± 0.10 | 1.21 ± 0.07 | 0.88 ± 0.04 |
| 2B12 | H | S | F | 2.63 ± 0.19 | 3.36 ± 0.26 | 1.05 ± 0.10 | 0.95 ± 0.03 | 0.92 ± 0.05 |
| 1G2 | R | R | F | 3.57 ± 0.21 | 2.51 ± 0.17 | 4.68 ± 0.13 | 1.77 ± 0.12 | 2.04 ± 0.18 |
| 2H11 | R | R | Y | 4.67 ± 0.24 | 2.65 ± 0.12 | 5.17 ± 0.49 | 1.87 ± 0.03 | 1.47 ± 0.13 |
| 3H5 | R | R | W | 3.78 ± 0.17 | 1.91 ± 0.12 | 4.09 ± 0.35 | 1.95 ± 0.13 | 2.06 ± 0.15 |
| 2A5 | Q | K | F | 8.06 ± 0.39 | 0.68 ± 0.09 | 2.12 ± 0.16 | 2.86 ± 0.19 | 4.00 ± 0.19 |
| 5F11 | Q | K | W | 8.45 ± 0.34 | 0.91 ± 0.06 | 2.31 ± 0.15 | 3.14 ± 0.21 | 2.97 ± 0.10 |
| 3G5 | Q | K | Y | 9.70 ± 0.60 | 0.59 ± 0.05 | 2.00 ± 0.11 | 2.47 ± 0.18 | 2.93 ± 0.08 |
| 3E7 | Q | R | W | 11.50 ± 0.83 | 1.31 ± 0.08 | 1.62 ± 0.17 | 3.98 ± 0.13 | 2.72 ± 0.17 |
| 1B1 | R | H | F | 2.33 ± 0.17 | 12.94 ± 0.69 | 7.32 ± 0.46 | 1.92 ± 0.08 | 1.65 ± 0.02 |
| 2E11 | R | H | Y | 2.51 ± 0.12 | 11.62 ± 0.21 | 6.29 ± 0.17 | 1.75 ± 0.02 | 1.34 ± 0.10 |
| 3C5 | R | L | F | 2.27 ± 0.13 | 9.73 ± 0.35 | 5.49 ± 0.37 | 3.05 ± 0.03 | 3.42 ± 0.15 |
| 4E1 | R | L | Y | 2.21 ± 0.06 | 9.70 ± 0.18 | 4.34 ± 0.18 | 2.78 ± 0.03 | 2.47 ± 0.14 |
Reactions were performed in Sörensen buffer employing 2 U mL−1 of each enzyme. The products were analyzed by HPAEC-PAD after approximately 90% sucrose was converted (reached in a range of 20–28 h reaction for all variants). Mean values and standard deviations of three independent experiments are shown. Mutated residues are shown as underlined bold letters.
aFor this sugar the peak relative area regarding to that of the wild type enzyme (set to 1.00) is provided.