Extended-spectrum class C β-lactamases have evolved to rapidly inactivate expanded-spectrum cephalosporins, a class of antibiotics designed to be resistant to hydrolysis by β-lactamase enzymes. To better understand the mechanism by which Acinetobacter-derived cephalosporinase-7 (ADC-7), a chromosomal AmpC enzyme, hydrolyzes these molecules, we determined the X-ray crystal structure of ADC-7 in an acyl-enzyme complex with the cephalosporin ceftazidime (2.40 Å) as well as in complex with a boronic acid transition state analog inhibitor that contains the R1 side chain of ceftazidime (1.67 Å). In the acyl-enzyme complex, the carbonyl oxygen is situated in the oxyanion hole where it makes key stabilizing interactions with the main chain nitrogens of Ser64 and Ser315. The boronic acid O1 hydroxyl group is similarly positioned in this area. Conserved residues Gln120 and Asn152 form hydrogen bonds with the amide group of the R1 side chain in both complexes. These complexes represent two steps in the hydrolysis of expanded-spectrum cephalosporins by ADC-7 and offer insight into the inhibition of ADC-7 by ceftazidime through displacement of the deacylating water molecule as well as blocking its trajectory to the acyl carbonyl carbon. In addition, the transition state analog inhibitor, LP06, was shown to bind with high affinity to ADC-7 (K_i, 50 nM) and was able to restore ceftazidime susceptibility, offering the potential for optimization efforts of this type of inhibitor.

中文翻译：

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头孢他啶及其硼酸过渡态类似物抑制不动杆菌衍生的头孢菌素酶ADC-7的结构见解

广谱C类β-内酰胺酶已经进化为可快速灭活广谱头孢菌素，广谱头孢菌素是一类旨在抵抗β-内酰胺酶水解的抗生素。为了更好地理解，通过该机制，一cinetobacter - d erived ÇEphalosporinase-7（ADC-7），一种染色体AmpC酶，水解这些分子，我们确定了带有头孢菌素头孢他啶（2.40Å）以及与包含头孢他啶R1侧链（1.67Å）的硼酸过渡态类似物抑制剂。在酰基酶复合物中，羰基氧位于氧阴离子孔中，与Ser64和Ser315的主链氮原子形成关键的稳定相互作用。硼酸O 1羟基类似地位于该区域中。保守残基Gln120和Asn152在两个复合物中均与R1侧链的酰胺基形成氢键。这些配合物代表了ADC-7水解广谱头孢菌素的两个步骤，并提供了头孢他啶通过脱酰基水分子的置换以及阻断其轨迹向酰基羰基碳的抑制作用。此外，已证明过渡态类似物抑制剂LP06与ADC-7（K _i（50 nM），并且能够恢复头孢他啶的药敏性，为优化这类抑制剂的潜力提供了可能。