FROM THE ENVIRONMENT TO THE CLINIC: KINETIC CHARACTERIZATION OF A CHROMOSOME-ENCODED PER-LIKE Β-LACTAMASE FROM THE ENVIRONMENTAL ISOLATE RHEINHEIMERA SP. KL1

RUGGIERO, MELINA; MUÑOZ, JACKSON IVAN BRICEÑO; RICCI, MILAGROS; POZZI, LUCIANA; GUTKIND, GABRIEL; POWER, PABLO

CABA

Congreso; VI INTERNATIONAL CONGRESS IN TRANSLATIONAL MEDICINE; 2023

The International Master in Biomedical Sciences (IMBS) and the Milstein-Köhler Binational PhD Program

β-lactamases are the main resistance mechanism to β-lactams in Gram negative bacteria. The β-lactamase encoding genes may be located on both plasmids or other mobile genetic elements, as well as ubiquitous genes in the chromosome of many environmental species from where they are recruited and transferred to pathogenic bacteria. PER enzymes belong to a family of class A serine-β-lactamases, with distinctive biochemical and structural features compared to other class A enzymes: i.e. high catalytic efficiency on all oxyimino-cephalosporins including both cefotaxime and ceftazidime, and differential structural features such as an enlarged and inverted omega-loop fold. PER-2 is a plasmid-encoded extended-spectrum β-lactamase (ESBL) present in clinical isolates of Enterobacterales in Argentina. The reservoir and origin of this family of β-lactamases is thought to be the chromosome of different environmental species of the proposed genus Pararheinheimera, recently splitted from the genus Rheinheimera. In order to determine the biochemical properties of PER enzymes encoded by these environmental species, the blaPER gene from Rheinheimera sp. KL1 (SAMN03447206) was cloned into a pET28a vector, the β-lactamase expressed and purified by affinity chromatoghrapgy (Ni Sepharose His-Trap). Steady-state kinetic parameters (kcat, Km, kcat/Km) were determined and compared with those of PER-2. The results indicate that this enzyme, named PER-KL1, has an ESBL profile and hydrolyze cephalothin more efficiently than PER-2 (kcat/Km relative to PER-2: 269%), although the hydrolysis of the rest of the tested antibiotics was less efficient (kcat/Km relative to PER-2 for ampicillin: 50%, ceftriaxone: 78%, ceftazidime: 2%, cefepime: 13%, aztreonam: 29%). Upon proper expression conditions (e.g., a suitable promoter and/or intensive antibiotic usage), recruitment of the PER-KL1 encoding gene in pathogenic bacteria would imply the dissemination of a new ESBL within the PER family. It remains to be determined how this enzyme will behave against classic β-lactamase inhibitors, diazabicyclooctanes inhibitors and the new siderophore cephalosporin cefiderocol.