Figure 2.

Tyr56 Is Critical for Stability of the Highly Conserved XTBD–XRN2ΔZLC Interface. (a) Surface representation of XRN2Δ^ZLC and XTBD structures highlighting conservation of the protein – protein interface (XTBD rotated 180° with respect to XRN2Δ^ZLC). Position-specific conservation scores computed by Consurf³³,³⁴ are displayed from white (weak to average conservation, Consurf levels 1-6) to red (most conserved, level 9). The dashed-dotted line on XRN2Δ^ZLC represents the projection of bound XTBD. Dashed lines on XTBD indicate residues analyzed in detail. (b-d) Representation of XTBD residues Cys54 and Tyr56 and their interaction with XRN2Δ^ZLC residues, zooming in on (c) interactions of the critical Tyr56 and (d) the side chain arrangement of Cys54 and Tyr56. Hydrogen bonds are indicated by gray dotted lines. (e) Gel showing results of Ni-NTA-mediated pull-down of His₆-PAXT-1 and its point mutant variants, respectively, co-expressed with XRN2. (f) Stereomicroscropic images of C. elegans grown as indicated schematically. The paxt-1(xe29) allele encodes PAXT-1_Y56A and was obtained by genome editing of endogenous paxt-1. (g) Western blot of 100 µg lysate from worms of the indicated genotype grown at 26°C using antibodies specific to the indicated endogenous proteins.