[Show abstract][Hide abstract] 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.
β-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.
[Show abstract][Hide abstract] ABSTRACT: Penicillin-binding protein (PBP) 2B from Streptococcus pneumoniae catalyzes the cross-linking of peptidoglycan precursors that occurs during bacterial cell-wall biosynthesis. A selenomethionyl (SeMet) substituted PBP 2B transpeptidase domain was isolated from a limited proteolysis digest of a soluble form of recombinant PBP 2B and then crystallized. The crystals belonged to space group P4(3)2(1)2, with unit-cell parameters a = b = 86.39, c = 143.27 A. Diffraction data were collected to 2.4 A resolution using the BL32B2 beamline at SPring-8. The asymmetric unit contains one protein molecule and 63.7% solvent.
Preview · Article · May 2008 · Acta Crystallographica Section F Structural Biology and Crystallization Communications
[Show abstract][Hide abstract] ABSTRACT: Enoyl-acyl carrier protein (ACP) reductases are critical for bacterial type II fatty acid biosynthesis and thus are attractive targets for developing novel antibiotics. We determined the crystal structure of enoyl-ACP reductase (FabK) from Streptococcus pneumoniae at 1.7 A resolution. There was one dimer per asymmetric unit. Each subunit formed a triose phosphate isomerase (TIM) barrel structure, and flavin mononucleotide (FMN) was bound as a cofactor in the active site. The overall structure was similar to the enoyl-ACP reductase (ER) of fungal fatty acid synthase and to 2-nitropropane dioxygenase (2-ND) from Pseudomonas aeruginosa, although there were some differences among these structures. We determined the crystal structure of FabK in complex with a phenylimidazole derivative inhibitor to envision the binding site interactions. The crystal structure reveals that the inhibitor binds to a hydrophobic pocket in the active site of FabK, and this is accompanied by induced-fit movements of two loop regions. The thiazole ring and part of the ureido moiety of the inhibitor are involved in a face-to-face pi-pi stacking interaction with the isoalloxazine ring of FMN. The side-chain conformation of the proposed catalytic residue, His144, changes upon complex formation. Lineweaver-Burk plots indicate that the inhibitor binds competitively with respect to NADH, and uncompetitively with respect to crotonoyl coenzyme A. We propose that the primary basis of the inhibitory activity is competition with NADH for binding to FabK, which is the first step of the two-step ping-pong catalytic mechanism.
[Show abstract][Hide abstract] ABSTRACT: Cefditoren is the active form of cefditoren pivoxil, an oral cephalosporin antibiotic used for the treatment of respiratory
tract infections and otitis media caused by bacteria such as Streptococcus pneumoniae, Haemophilus influenzae, Streptococcus pyogenes, Klebsiella pneumoniae, and methicillin-susceptible strains of Staphylococcus aureus. β-Lactam antibiotics, including cefditoren, target penicillin-binding proteins (PBPs), which are membrane-associated enzymes
that play essential roles in the peptidoglycan biosynthetic process. To envision the binding of cefditoren to PBPs, we determined
the crystal structure of a trypsin-digested form of PBP 2X from S. pneumoniae strain R6 complexed with cefditoren. There are two PBP 2X molecules (designated molecules 1 and 2) per asymmetric unit. The
structure reveals that the orientation of Trp374 in each molecule changes in a different way upon the formation of the complex,
but each forms a hydrophobic pocket. The methylthiazole group of the C-3 side chain of cefditoren fits into this binding pocket,
which consists of residues His394, Trp374, and Thr526 in molecule 1 and residues His394, Asp375, and Thr526 in molecule 2.
The formation of the complex is also accompanied by an induced-fit conformational change of the enzyme in the pocket to which
the C-7 side chain of cefditoren binds. These features likely play a role in the high level of activity of cefditoren against
Preview · Article · Dec 2007 · Antimicrobial Agents and Chemotherapy
[Show abstract][Hide abstract] ABSTRACT: The enoyl-acyl carrier protein (ACP) reductase from Streptococcus pneumoniae (FabK; EC 220.127.116.11) is responsible for catalyzing the final step in each elongation cycle of fatty-acid biosynthesis. Selenomethionine-substituted FabK was purified and crystallized by the hanging-drop vapour-diffusion method at 277 K. The crystal belongs to space group P2(1), with unit-cell parameters a = 50.26, b = 126.70, c = 53.63 A, beta = 112.46 degrees . Diffraction data were collected to 2.00 A resolution using synchrotron beamline BL32B2 at SPring-8. Two molecules were estimated to be present in the asymmetric unit, with a solvent content of 45.1%.
Preview · Article · Jul 2006 · Acta Crystallographica Section F Structural Biology and Crystallization Communications