Table 1. Double phosphorylation of Pto peptide 187–202 and 188–202 upon activation of signalling.
| Prf | prfK1128A | prfD1416V | prf1–546 | ||||
| Pto peptide 188–202 | EV* (3 dpi) | AvrPto (2 dpi) | AvrPtoB (2 dpi) | EV (2 dpi) control | EV (2 dpi) | EV (2 dpi) | EV (2 dpi) |
| G[pT190]ELDQTHLSTVVK | 0 | 0 | 0 | 0 | 0 | 0 | 1 |
| GTELDQTHL[pS198]TVVK | 2 | 4 | 3 | 9 | 16 | 13 | 20 |
| GTELDQTHLS [pT199]VVK | 2 | 3 | 3 | 8 | 3 | 3 | 7 |
| GTELDQTHL[pS198][pT199]VVK | 2 | 7 | 2 | 0 | 0 | 1 | 0 |
| GTELDQTHLSTVVK | 21 | 20 | 20 | 33 | 48 | 6 | 2 |
| Pto peptide 187–202 | |||||||
| KGTELDQ[pT195]HLS TVVK | 0 | 0 | 0 | 0 | 0 | 1 | 0 |
| KGTELDQTHL[pS198]TVVK | 14 | 14 | 11 | 14 | 16 | 8 | 5 |
| KGTELDQTHLS [pT199]VVK | 0 | 2 | 0 | 4 | 3 | 1 | 1 |
| KG[pT190]ELDQTHLS[pT199]VVK | 1 | 0 | 0 | 0 | 0 | 1 | 0 |
| KGTELDQ[pT195]HLS [pT199]VVK | 0 | 1 | 0 | 0 | 0 | 0 | 0 |
| KGTELDQTHL[pS198][pT199]VVK | 1 | 5 | 3 | 0 | 0 | 4 | 0 |
| KGTELDQTHLSTVVK | 11 | 32 | 8 | 14 | 26 | 6 | 5 |
| peptides 187–202 and 188–202 | 54 | 88 | 50 | 83 | 112 | 47 | 41 |
| percentage of peptides with [pS198] and [pT199] | 5.5% | 13.6% | 10% | 0% | 0% | 10.6% | 0% |
| Pto Sequence Coverage | 87% | 75% | 82% | 76% | 79% | 38% | 53% |
| Signalling (HR) | yes * | yes | yes | no | no | yes | no |
Prf-3HA, prfK1128A –3HA, prfD1416V –3HA, N-term -3HA (prf1–546 –3HA), Pto-FLAG AvrPto and AvrPtoB were expressed transiently in N. benthamiana under the control of 35S promoter; the total amount of Pto-FLAG was immunoprecipitated using anti-FLAG antibodies. The number of peptides identified with 0, 1, and 2 phosphorylation events is indicated.
*
ligand-independent HR.