Crystal structures of biapenem and tebipenem complexed with penicillin-binding proteins 2X and 1A from Streptococcus pneumoniae

Abstract

Biapenem is a parenteral carbapenem antibiotic that exhibits wide-ranging antibacterial activity, remarkable chemical stability, and extensive stability against human renal dehydropeptidase-I. Tebipenem is the active form of tebipenem pivoxil, a novel oral carbapenem antibiotic that has a high level of bioavailability in humans, in addition to the above-mentioned features. beta-lactam antibiotics, including carbapenems, target penicillin-binding proteins (PBPs), which are membrane-associated enzymes that play essential roles in peptidoglycan biosynthesis. To envisage the binding of carbapenems to PBPs, we determined the crystal structures of the trypsin-digested forms of both PBP 2X and PBP 1A from Streptococcus pneumoniae strain R6, each complexed with biapenem or tebipenem. The structures of the complexes revealed that the carbapenem C-2 side chains form hydrophobic interactions with Trp374 and Thr526 of PBP 2X and with Trp411 and Thr543 of PBP 1A. The Trp and Thr residues are conserved in PBP 2B. These results suggest that interactions between the C-2 side chains of carbapenems and the conserved Trp and Thr residues in PBPs play important roles in the binding of carbapenems to PBPs.

FIG. 1.

Chemical structures of biapenem and tebipenem, with the atomic numbering scheme of the carbapenem skeleton.

FIG. 2.

PBP 2X-carbapenem complex structures. (A and B) F_o − F_c omit electron density maps calculated for the structures omitting the carbapenem moiety covalently attached to Ser337 in molecule 1 of PBP 2X in complex with biapenem and tebipenem. The maps, depicted as magenta meshes, are contoured at 2.8 σ. Carbapenem-acylated Ser residues are shown as stick models, with carbon atoms colored green, oxygen atoms red, nitrogen atoms blue, and sulfur atoms yellow. (C) Superpositioned active-site regions of the uncomplexed (magenta) and biapenem (light blue)- and tebipenem (orange)-complexed structures. The carbapenems and side chains of selected residues are shown as thick sticks. Atoms are colored as described for panel A, except that the carbon atoms of the carbapenems and selected residues in the uncomplexed and complexed structures are magenta, light blue, and orange, respectively. (D and E) Stereo view of the PBP 2X active site complexed with biapenem and tebipenem, respectively. Atoms are colored as described for panel A, except that the carbon atoms of PBP 2X are orange. The positions of hydrogen atoms of the C-2 side chain were calculated by DS Modeling (Accelrys). The hydrogen atoms that form CH/π interactions are shown as balls (light blue), and other hydrogen atoms are shown as thick sticks (white). Hydrogen bonds are represented by thin lines (magenta). (F) Schematic diagram of the carbapenem binding mode. Hydrogen bonds are represented by broken lines (magenta), and hydrophobic contacts are shown as arcs (green). The atomic numbering scheme of the carbapenem skeleton is also shown.

FIG. 3.

PBP1A-carbapenem complex structures. (A and B) F_o − F_c omit electron density maps calculated for the structures omitting the carbapenem moiety covalently attached to Ser370 in molecule 1 of PBP 1A in complex with biapenem and tebipenem. The maps and models are displayed in the same manner as for Fig. 2A and B. (C) Superpositioned active-site regions of the uncomplexed (magenta) and biapenem (light blue)- and tebipenem (orange)-complexed structures. The carbapenems and side chains of selected residues are shown as thick sticks. Atoms are colored as described for Fig. 2C. (D and E) Stereo view of the active sites of PBP 1A in complex with biapenem and tebipenem, respectively. Atoms are colored as described for Fig. 2A and B, except that the carbon atoms of PBP 1A are orange. Hydrogen atoms of the C-2 side chain and hydrogen bonds are displayed in the same manner as for Fig. 2D and E. (F) Schematic diagram of the carbapenem binding mode. Hydrogen bonds are represented by broken lines (magenta), and hydrophobic contacts are shown as arcs (green). The atomic numbering scheme of the carbapenem skeleton is also shown.

FIG. 4.

Comparison of the PBP 2X and PBP 1A complexes. (A) Superpositioned active-site regions of PBP 2X (light blue)- and PBP 1A (orange)-biapenem complex structures. Biapenem and the side chains of the Trp and Thr residues in the active sites are shown as thick sticks. Atoms are colored as described for Fig. 2A, except that the carbon atoms in PBP 2X- and PBP 1A-carbapenem complex structures are light blue and orange, respectively. (B) Superpositioned active-site regions of PBP 2X (light blue)- and PBP 1A (orange)-tebipenem complex structures. The models are displayed in the same manner as for panel A.

FIG. 5.

(A)Chemical structures of compound 1, a cephalosporin that mimics the acceptor strand of a Streptomyces sp. strain R61 peptidoglycan. A diaminopimelate surrogate is shown in red. (B) Superpositioned active-site regions of the S. pneumoniae PBP 2X-biapenem complex (light blue) and the Streptomyces dd-peptidase/transpeptidase-compound 1 complex (orange) structures. Biapenem, the side chains of the Trp 374 and Thr526 residues, and compound 1 are shown as thick sticks. Atoms are colored as described for Fig. 2A, except that the carbon atoms in PBP 2X and the dd-peptidase/transpeptidase complex structures are light blue and orange, respectively.

FIG. 6.

Chemical structures of compound 2, compound 3, penicillin G, and cephalothin, with the atomic numbering scheme of the penicillin and cephalosporin skeleton.