Extended Data Fig. 3: Sequence alignment of the SBC regions in NPR proteins from several plant species, and details of the SA-binding pocket and activity.
From: Structural basis of salicylic acid perception by Arabidopsis NPR proteins
Related to Fig. 2. a, Structure-based sequence alignment of the SBC regions of NPR4 and NPR1 orthologues. The secondary-structure diagram of NPR4 SBC is shown above the sequences. Regions with no regular secondary structure are shown by lines, and α-helices are represented by cylinders. The dashed lines indicate two disordered loops that are not resolved in the structure. Strictly conserved residues are coloured in blue. The rest of the residues are coloured with black (87.5%), brown (75%) or red (<75%) based on their degrees of conservation. The residues directly involved in SA binding are highlighted with asterisks. The putative EAR motif is labelled and indicated by a black bar. Six surface residues selected for mutagenesis analysis are labelled. At (Arabidopsis thaliana): NPR1 AT1G64280, NPR3 AT5G45110 and NPR4 AT4G19660) Os (Oryza sativa): NH1 Os01g09800, NH2 Os01g56200 and NH3 Os03g46440; Nb (Nicotiana benthamiana): NPR1 LOC107831756; Bn (Brassica napa) NPR1 LOC106389246. b, A close-up stereo view of the NPR4 SBC SA-binding pocket with the omit map electron density, shown together with the residues in the stick model. SA is coloured in yellow and red and situated in the centre. Three selected SA-contacting residues in close proximity to the SA carboxyl group are indicated. c, Ligplot of the hydrophobic and polar interactions between SA and NPR4-SBC residues. d, A semi-transparent view of the SA-binding pocket with the SA analogue benzothiadiazole (BTH) (magenta, blue and red sticks) modelled onto SA (yellow and red sticks) situated in the centre and indicated by arrows. The view is related to the NPR4 SBC internal cavity shown in Fig. 2c by 180° vertical rotation. Ala434 is shown as a yellow stick, and indicated as A434. The internal cavity and surrounding surfaces of NPR4 SBC are shown in green surface representation. e, SA binding by wild-type NPR3 (WT) and NPR3(R428A) as determined with radiolabelled ligand binding assay with 100 nM 3H-SA. n = 6 independent samples. Error bars indicate s.d. (centre value).
