High-Throughput Crystallography Reveals Boron-Containing Inhibitors of a Penicillin-Binding Protein with Di- and Tricovalent Binding Modes

The effectiveness
of β-lactam antibiotics is increasingly
compromised by β-lactamases. Boron-containing inhibitors are
potent serine-β-lactamase inhibitors, but the interactions of
boron-based compounds with the penicillin-binding protein (PBP) β-lactam
targets have not been extensively studied. We used high-throughput
X-ray crystallography to explore reactions of a boron-containing fragment
set with the Pseudomonas aeruginosa PBP3 (PaPBP3). Multiple crystal structures reveal that boronic acids
react with PBPs to give tricovalently linked complexes bonded to Ser294,
Ser349, and Lys484 of PaPBP3; benzoxaboroles react with PaPBP3 via
reaction with two nucleophilic serines (Ser294 and Ser349) to give
dicovalently linked complexes; and vaborbactam reacts to give a monocovalently
linked complex. Modifications of the benzoxaborole scaffold resulted
in a moderately potent inhibition of PaPBP3, though no antibacterial
activity was observed. Overall, the results further evidence the potential
for the development of new classes of boron-based antibiotics, which
are not compromised by β-lactamase-driven resistance.

β-内酰胺类抗生素的抗菌效力正日益受到β-内酰胺酶的削弱。含硼抑制剂是一类强效的丝氨酸β-内酰胺酶抑制剂，但目前针对硼基化合物与青霉素结合蛋白（penicillin-binding protein, PBP）这类β-内酰胺作用靶点的相互作用研究仍不够充分。本研究采用高通量X射线晶体学技术，探究了含硼片段库与铜绿假单胞菌PBP3（PaPBP3）的反应过程。多组晶体结构解析结果表明：硼酸类化合物可与PBP结合，形成与PaPBP3的Ser294、Ser349及Lys484残基以三共价键结合的复合物；苯并氧硼杂环类化合物则通过与两个亲核丝氨酸残基（Ser294与Ser349）发生反应，生成双共价键结合的复合物；而阿维巴坦（vaborbactam）仅能与PaPBP3形成单共价键结合的复合物。对苯并氧硼杂环类母核进行结构修饰后，可获得对PaPBP3具有中等强度抑制活性的化合物，但未检测到抗菌活性。综上，本研究结果进一步证实了开发新型硼基抗生素的潜力，这类抗生素不会受到β-内酰胺酶介导的耐药性影响。