Fig. 3: Architectural organization of plant CYP73A5^Δ2-28 and ATR2^Δ2-77 based on self-assembled peptide bio-machinery in E. coli.

a The peptide ligation system SpySystem, spontaneously forming a post-translational intermolecular isopeptide bond (magenta) between the side chains of Lys34 in SpyCatcher peptide (gray) and Asp7 in SpyTag peptide (black)³⁰, was harnessed to serve as a self-folded dual-track bridge to organize CYP73A5^Δ2-28 and ATR2^Δ2-77. When SpyCatcher and SpyTag were respectively fused to the C-terminus of CYP73A5^Δ2-28 and the N-terminus of ATR2^Δ2-77, the covalent heterodimer (I) formed post-translationally. When SpyCatcher and SpyTag were respectively fused to the N-terminus of CYP73A5^Δ2-28 and the C-terminus of ATR2^Δ2-77, the covalent heterodimer (II) formed post-translationally. When SpyCatcher and SpyTag were respectively fused to the C-termini of CYP73A5^Δ2-28 and ATR2^Δ2-77, the covalent heterodimer (III) formed post-translationally. When SpyCatcher and SpyTag were respectively fused to the N-termini of CYP73A5^Δ2-28 and ATR2^Δ2-77, the covalent heterodimer (IV) formed post-translationally. The arrow indicates a polypeptide from the N-terminus to the C-terminus. b p-Coumaric acid de novo biosynthesis (upper panel) and trans-cinnamic acid accumulation (lower panel) of the AthPAL1-expressing E. coli strains with the heterodimer (I), (II), (III) or (IV). Data are shown as mean ± SE (n = 11 (I), 10 (II), 12 (III), 12 (IV) biological independent clones). Significant differences (P < 0.05) of p-coumaric acid titer at 48 hpi are evaluated by one-way ANOVA with Duncan’s new multiple range test and indicated by different lowercase letters. Source data are provided as a Source Data file.