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Biochemical and structural analysis of inhibitors targeting the ADC-7 cephalosporinase of Acinetobacter baumannii.

Powers RA, Swanson HC, Taracila MA, Florek NW, Romagnoli C, Caselli E, Prati F, Bonomo RA, Wallar BJ - Biochemistry (2014)

Bottom Line: Currently, β-lactamase inhibitors are structurally similar to β-lactam substrates and are not effective inactivators of this class C cephalosporinase.In addition, the carboxylate group of the inhibitor is meant to mimic the C3/C4 carboxylate found in β-lactams.The ADC-7/BATSI complex provides insight into recognition of non-β-lactam inhibitors by ADC enzymes and offers a starting point for the structure-based optimization of this class of novel β-lactamase inhibitors against a key resistance target.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Grand Valley State University , 1 Campus Drive, Allendale, Michigan 49401, United States.

ABSTRACT
β-Lactam resistance in Acinetobacter baumannii presents one of the greatest challenges to contemporary antimicrobial chemotherapy. Much of this resistance to cephalosporins derives from the expression of the class C β-lactamase enzymes, known as Acinetobacter-derived cephalosporinases (ADCs). Currently, β-lactamase inhibitors are structurally similar to β-lactam substrates and are not effective inactivators of this class C cephalosporinase. Herein, two boronic acid transition state inhibitors (BATSIs S02030 and SM23) that are chemically distinct from β-lactams were designed and tested for inhibition of ADC enzymes. BATSIs SM23 and S02030 bind with high affinity to ADC-7, a chromosomal cephalosporinase from Acinetobacter baumannii (Ki = 21.1 ± 1.9 nM and 44.5 ± 2.2 nM, respectively). The X-ray crystal structures of ADC-7 were determined in both the apo form (1.73 Å resolution) and in complex with S02030 (2.0 Å resolution). In the complex, S02030 makes several canonical interactions: the O1 oxygen of S02030 is bound in the oxyanion hole, and the R1 amide group makes key interactions with conserved residues Asn152 and Gln120. In addition, the carboxylate group of the inhibitor is meant to mimic the C3/C4 carboxylate found in β-lactams. The C3/C4 carboxylate recognition site in class C enzymes is comprised of Asn346 and Arg349 (AmpC numbering), and these residues are conserved in ADC-7. Interestingly, in the ADC-7/S02030 complex, the inhibitor carboxylate group is observed to interact with Arg340, a residue that distinguishes ADC-7 from the related class C enzyme AmpC. A thermodynamic analysis suggests that ΔH driven compounds may be optimized to generate new lead agents. The ADC-7/BATSI complex provides insight into recognition of non-β-lactam inhibitors by ADC enzymes and offers a starting point for the structure-based optimization of this class of novel β-lactamase inhibitors against a key resistance target.

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Superposition of ADC-7/S02030(salmon) complex with AmpC/SM23 (white). Boronic acid inhibitor (SM23)is bound to the AmpC active site and highlights the position of theinhibitor’s carboxylate group that is meant to mimic the correspondinggroup common to β-lactams.
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fig5: Superposition of ADC-7/S02030(salmon) complex with AmpC/SM23 (white). Boronic acid inhibitor (SM23)is bound to the AmpC active site and highlights the position of theinhibitor’s carboxylate group that is meant to mimic the correspondinggroup common to β-lactams.

Mentions: The next step is to improvethe properties of S02030 so that the high affinity also translatesinto increased activity against resistant bacteria. Unlike the achiralcephalothin BATSI from which they are derived, SM23 and S02030 containboth an R1 and an R2 group (Figure 1C). Thesetwo regions are important for exploring different areas of the activesite, as well as for identifying important inhibitor recognition sitesto aid in future optimization efforts. Focusing first on the R2 group,the ADC-7/S02030 complex showed differences in S02030 binding, coupledwith multiple Arg340 conformations (Figure 4) in each of the four monomers of ADC-7. This lack of a single consensusbinding mode for the inhibitor suggests that the R2 carboxylate groupof S02030 may be too close to Arg340, resulting in a steric clashbetween this residue and the carboxylate group of the inhibitor. Fromthe AmpC/SM23 (PDB 1MXO) comparison, removal of the carbon linker may place the carboxylatein an ideal position for a favorable interaction with Arg340 (Figure 5). The shorter R2 group of SM23 may allow for afavorable and stabilizing interaction with Arg340 and may explainwhy SM23 has a higher affinity for ADC-7.


Biochemical and structural analysis of inhibitors targeting the ADC-7 cephalosporinase of Acinetobacter baumannii.

Powers RA, Swanson HC, Taracila MA, Florek NW, Romagnoli C, Caselli E, Prati F, Bonomo RA, Wallar BJ - Biochemistry (2014)

Superposition of ADC-7/S02030(salmon) complex with AmpC/SM23 (white). Boronic acid inhibitor (SM23)is bound to the AmpC active site and highlights the position of theinhibitor’s carboxylate group that is meant to mimic the correspondinggroup common to β-lactams.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4263437&req=5

fig5: Superposition of ADC-7/S02030(salmon) complex with AmpC/SM23 (white). Boronic acid inhibitor (SM23)is bound to the AmpC active site and highlights the position of theinhibitor’s carboxylate group that is meant to mimic the correspondinggroup common to β-lactams.
Mentions: The next step is to improvethe properties of S02030 so that the high affinity also translatesinto increased activity against resistant bacteria. Unlike the achiralcephalothin BATSI from which they are derived, SM23 and S02030 containboth an R1 and an R2 group (Figure 1C). Thesetwo regions are important for exploring different areas of the activesite, as well as for identifying important inhibitor recognition sitesto aid in future optimization efforts. Focusing first on the R2 group,the ADC-7/S02030 complex showed differences in S02030 binding, coupledwith multiple Arg340 conformations (Figure 4) in each of the four monomers of ADC-7. This lack of a single consensusbinding mode for the inhibitor suggests that the R2 carboxylate groupof S02030 may be too close to Arg340, resulting in a steric clashbetween this residue and the carboxylate group of the inhibitor. Fromthe AmpC/SM23 (PDB 1MXO) comparison, removal of the carbon linker may place the carboxylatein an ideal position for a favorable interaction with Arg340 (Figure 5). The shorter R2 group of SM23 may allow for afavorable and stabilizing interaction with Arg340 and may explainwhy SM23 has a higher affinity for ADC-7.

Bottom Line: Currently, β-lactamase inhibitors are structurally similar to β-lactam substrates and are not effective inactivators of this class C cephalosporinase.In addition, the carboxylate group of the inhibitor is meant to mimic the C3/C4 carboxylate found in β-lactams.The ADC-7/BATSI complex provides insight into recognition of non-β-lactam inhibitors by ADC enzymes and offers a starting point for the structure-based optimization of this class of novel β-lactamase inhibitors against a key resistance target.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry, Grand Valley State University , 1 Campus Drive, Allendale, Michigan 49401, United States.

ABSTRACT
β-Lactam resistance in Acinetobacter baumannii presents one of the greatest challenges to contemporary antimicrobial chemotherapy. Much of this resistance to cephalosporins derives from the expression of the class C β-lactamase enzymes, known as Acinetobacter-derived cephalosporinases (ADCs). Currently, β-lactamase inhibitors are structurally similar to β-lactam substrates and are not effective inactivators of this class C cephalosporinase. Herein, two boronic acid transition state inhibitors (BATSIs S02030 and SM23) that are chemically distinct from β-lactams were designed and tested for inhibition of ADC enzymes. BATSIs SM23 and S02030 bind with high affinity to ADC-7, a chromosomal cephalosporinase from Acinetobacter baumannii (Ki = 21.1 ± 1.9 nM and 44.5 ± 2.2 nM, respectively). The X-ray crystal structures of ADC-7 were determined in both the apo form (1.73 Å resolution) and in complex with S02030 (2.0 Å resolution). In the complex, S02030 makes several canonical interactions: the O1 oxygen of S02030 is bound in the oxyanion hole, and the R1 amide group makes key interactions with conserved residues Asn152 and Gln120. In addition, the carboxylate group of the inhibitor is meant to mimic the C3/C4 carboxylate found in β-lactams. The C3/C4 carboxylate recognition site in class C enzymes is comprised of Asn346 and Arg349 (AmpC numbering), and these residues are conserved in ADC-7. Interestingly, in the ADC-7/S02030 complex, the inhibitor carboxylate group is observed to interact with Arg340, a residue that distinguishes ADC-7 from the related class C enzyme AmpC. A thermodynamic analysis suggests that ΔH driven compounds may be optimized to generate new lead agents. The ADC-7/BATSI complex provides insight into recognition of non-β-lactam inhibitors by ADC enzymes and offers a starting point for the structure-based optimization of this class of novel β-lactamase inhibitors against a key resistance target.

Show MeSH