TheEscherichia coli(E. coli) ribosome can incorporate a variety of non-l-α-amino
acid monomers into polypeptide chainsin vitrobut
with poor efficiency. Although these monomers span a diverse set of
compounds, there exists no high-resolution structural information
regarding their positioning within the catalytic center of the ribosome,
the peptidyl transferase center (PTC). Thus, details regarding the
mechanism of amide bond formation and the structural basis for differences
and defects in incorporation efficiency remain unknown. Within a set
of three aminobenzoic acid derivatives—3-aminopyridine-4-carboxylic
acid (Apy),ortho-aminobenzoic acid (oABZ), andmeta-aminobenzoic acid (mABZ)—the ribosome incorporates Apy into polypeptide chains
with the highest efficiency, followed byoABZ and
thenmABZ, a trend that does not track with the nucleophilicity
of the reactive amines. Here, we report high-resolution cryo-EM structures
of the ribosome with each of these three aminobenzoic acid derivatives
charged on tRNA bound in the aminoacyl-tRNA site (A-site). The structures
reveal how the aromatic ring of each monomer sterically blocks the
positioning of nucleotide U2506, thereby preventing rearrangement
of nucleotide U2585 and the resulting induced fit in the PTC required
for efficient amide bond formation. They also reveal disruptions to
the bound water network that is believed to facilitate formation and
breakdown of the tetrahedral intermediate. Together, the cryo-EM structures
reported here provide a mechanistic rationale for differences in reactivity
of aminobenzoic acid derivatives relative tol-α-amino
acids and each other and identify stereochemical constraints on the
size and geometry of non-monomers that can be accepted efficiently
by wild-type ribosomes.