Table 1.
Spectral and Kinetic Constants of Truncation and Point Mutants of Arabidopsis PhyB.
| Mutationa | Absorption maximab (nm) | Photoconversion ratec,d,e (min−1) | Therm revc,d,f (x1000 min−1) | ||
|---|---|---|---|---|---|
| Pr | Pfr | Pr ⇒ Pfr | Pfr ⇒ Pr | Pfr ⇒ Pr | |
| Domain Truncations | |||||
| PΦB | |||||
| 1–450 | 658 | 707 | 0.72g | 0.11 (±0.02) | 10.5 (±0.1) |
| 1–624 | 664 | 724 | 0.98 (±0.03) | 2.45 (±0.04) | 7.91 (±0.02) |
| 1–908h | 665 | 724 | k1 2.1 (±0.3) | 2.6 (±0.2) | 320 (±20) |
| k2 0.47 (±0.05) | 34 (±3) | ||||
| FLh,i | 664 | 725 | k1 1.1 | 410 | |
| k2 0.12 | 40 | ||||
| PCB | |||||
| 1–450 | 643 | 693 | 1.04 (±0.05) | 0.38 (±0.02) | 38.4 ( ± 0.1) |
| 1–624 | 651 | 714 | 1.35 (±0.04) | 3.52 (±0.03) | 0.52 (±0.01) |
| 1–908 | 651 | 714 | 1.20 (±0.08) | 3.52 (±0.06) | 29.9 (±0.3) |
| NTE Truncations | |||||
| 10–624 | 665 | 724 | 0.86 (±0.04) | 2.43 (±0.01) | 9.54 (±0.02) |
| 20–624 | 665 | 724 | 0.82 (±0.09) | 2.52 (±0.05) | 10.1 (±0.1) |
| 30–624 | 665 | 724 | 0.84 (±0.03) | 2.54 (±0.07) | 7.7 (±0.5) |
| 41–624 | 665 | 725 | 0.88 (±0.03) | 2.60 (±0.06) | 6.50 (±0.02) |
| 46–624 | 666 | 725 | 10.8 (±0.1) | ||
| 51–624 | 664 | 723 | 1.1 (±0.1) | 2.53 (±0.06) | 16.2 (±0.1) |
| 55–624 | 665 | 724 | 14.9 (±0.1) | ||
| 56–624 | 665 | 723 | 32.3 (±0.1) | ||
| 57–624 | 665 | 722 | 0.97 (±0.05) | 2.35 (±0.09) | 50.1 (±0.1) |
| 59–624 | 664 | 719 | 91.8 (±0.3) | ||
| 60–624 | 665 | 720 | 1.09 (±0.02) | 2.39 (±0.06) | 85 (±1) |
| 65–624 | 666 | 717 | 1.18 (±0.09) | 143 (±1) | |
| 70–624 | 665 | 717 | 1.05 (±0.06) | 2.28 (±0.03) | 146 (±1) |
| 80–624 | 664 | 717 | 1.17 (±0.03) | 2.3 (±0.1) | 169 (±1) |
| 90–624 | 664 | 717 | 1.3g | 2.5g | 163 (±1) |
| 100–624 | 664 | 717 | 0.99 (±0.05) | 2.2 (±0.2) | 134 (±1) |
| PSM Point Mutants | |||||
| S55A,K56A | 666 | 725 | 16.3 (±0.1) | ||
| Q59A,Q60A | 666 | 721 | 35.5 (±0.1) | ||
| Y61A,T62A | 665 | 725 | 7.53 (±0.01) | ||
aData were collected from PhyB containing the indicated residues after removal of the N-terminal 6His-TEV tag.
bMaxima of the Q absorption band. Pfr maxima were identified from difference spectra generated by subtracting red-irradiated from dark-adapted samples.
cCalculated from the global fit of absorption at 640 nm, 650 nm, 660 nm, 670 nm, 700 nm, 710 nm, 720 nm, 730 nm, and 740 nm as a function of time using the equation: Abs = ΔAbstotal•exp(−k•Time) + Absinitial.
dValues in parentheses represent the average of three technical replicates (±SD).
eFluence rate was calibrated to 12 μmol•m−2•s−1 at the intersecting spectrophotometric light path.
fFluence rate was calibrated to a nominal 0.465 μmol•m−2•s−1 for each measurement.
gAverage of two technical replicates.
hPr ⇒ Pfr photoconversion, Pfr ⇒ Pr photoconversion, and/or thermal reversion were expressed as the sum of two exponentials using the equation: Abs = ΔAbstotal1•exp(−k1*Time) + ΔAbstotal2•exp(−k2•Time) + Absinitial.
iData were collected from a single sample.