Extended Data Fig. 7: No higher oligomers were observed from PsnB.
From: Molecular mechanism underlying substrate recognition of the peptide macrocyclase PsnB
a, Gel-filtration chromatogram of PsnB without MgCl2 (black solid line), with MgCl2 (red solid line), or with both MgCl2 and ATP (blue solid line). A chromatogram of molecular weight standard (black dashed line) is also shown with known molecular weights. Addition of nucleotide did not induce the formation of the stable higher oligomer of PsnB dimer. b‒e, FRET of a solution containing both donor- and acceptor-labeled PsnB was measured with different amounts of MgCl2 (b), AMPPNP (c), LP (d), or MP (e). Additional components in solutions are listed above the plots. Neither nucleotide nor precursor induced stable intermolecular interaction of PsnB. f, Loop modeling revealed that β13β14 loop is long and flexible enough for intramolecular interaction. Residues between Ile227 and Ile247, the β13β14 loop region, were modeled by FALC59,60 and overall complex structure was optimized by relaxation. In the modeled structure, the conformation of the β13β14 loop was flipped and the DFR motif moved toward the enzyme active site. Also, Arg235 had hydrogen bonding with nucleotide which is similar to the intermolecular interaction scheme shown in crystal structures (Fig. 4c).
