TY - JOUR

T1 - Revisiting the hydrolysis of ampicillin catalyzed by Temoneira-1 β-lactamase, and the effect of Ni(II), Cd(II) and Hg(II)

AU - Nafaee, Zeyad H.

AU - Egyed, Viktória

AU - Jancsó, Attila

AU - Tóth, Annamária

AU - Gerami, Adeleh Mokhles

AU - Dang, Thanh Thien

AU - Heiniger-Schell, Juliana

AU - Hemmingsen, Lars

AU - Hunyadi-Gulyás, Éva

AU - Peintler, Gábor

AU - Gyurcsik, Béla

N1 - Funding Information: This research was supported by the Hungarian National Research, Development and Innovation Office (GINOP-2.3.2-15-2016-00038, GINOP-2.3.2-15-2016-00001, GINOP-2.3.2-15-2016-00020, and 2019-2.1111-TÉT-2019-00089). We acknowledge the financial support received from the Federal Ministry of Education and Research (BMBF) through grants 05K16PGA and 05K22PGA. We also thank the European Union's Horizon Europe Framework research and innovation programme under grant agreement no. 101057511 (EURO-LABS) and the European Union's Horizon 2020 Framework research and innovation program under grant agreement no. 654002 (ENSAR2). Funding project CERN-FIS-PAR-0005-2017 FCT-Portugal is acknowledged in support of the analogue PAC setup and of the new implantation chamber for icy samples. We thank CERN and the ISOLDE technical team for beam time, EURONS and NICE for financial support. Funding Information: This research was supported by the Hungarian National Research, Development and Innovation Office (GINOP‐2.3.2‐15‐2016‐00038, GINOP‐2.3.2‐15‐2016‐00001, GINOP‐2.3.2‐15‐2016‐00020, and 2019‐2.1111‐TÉT‐2019‐00089). We acknowledge the financial support received from the Federal Ministry of Education and Research (BMBF) through grants 05K16PGA and 05K22PGA. We also thank the European Union's Horizon Europe Framework research and innovation programme under grant agreement no. 101057511 (EURO‐LABS) and the European Union's Horizon 2020 Framework research and innovation program under grant agreement no. 654002 (ENSAR2). Funding project CERN‐FIS‐PAR‐0005‐2017 FCT‐Portugal is acknowledged in support of the analogue PAC setup and of the new implantation chamber for icy samples. We thank CERN and the ISOLDE technical team for beam time, EURONS and NICE for financial support. Publisher Copyright: © 2023 The Authors. Protein Science published by Wiley Periodicals LLC on behalf of The Protein Society.

PY - 2023

Y1 - 2023

N2 - β-Lactamases grant resistance to bacteria against β-lactam antibiotics. The active center of TEM-1 β-lactamase accommodates a Ser-Xaa-Xaa-Lys motif. TEM-1 β-lactamase is not a metalloenzyme but it possesses several putative metal ion binding sites. The sites composed of His residue pairs chelate borderline transition metal ions such as Ni(II). In addition, there are many sulfur-containing donor groups that can coordinate soft metal ions such as Hg(II). Cd(II) may bind to both types of the above listed donor groups. No significant change was observed in the circular dichroism spectra of TEM-1 β-lactamase on increasing the metal ion content of the samples, with the exception of Hg(II) inducing a small change in the secondary structure of the protein. A weak nonspecific binding of Hg(II) was proven by mass spectrometry and 119mHg perturbed angular correlation spectroscopy. The hydrolytic process of ampicillin catalyzed by TEM-1 β-lactamase was described by the kinetic analysis of the set of full catalytic progress curves, where the slow, yet observable conversion of the primary reaction product into a second one, identified as ampilloic acid by mass spectrometry, needed also to be considered in the applied model. Ni(II) and Cd(II) slightly promoted the catalytic activity of the enzyme while Hg(II) exerted a noticeable inhibitory effect. Hg(II) and Ni(II), applied at 10 μM concentration, inhibited the growth of E. coli BL21(DE3) in M9 minimal medium in the absence of ampicillin, but addition of the antibiotic could neutralize this toxic effect by complexing the metal ions.

AB - β-Lactamases grant resistance to bacteria against β-lactam antibiotics. The active center of TEM-1 β-lactamase accommodates a Ser-Xaa-Xaa-Lys motif. TEM-1 β-lactamase is not a metalloenzyme but it possesses several putative metal ion binding sites. The sites composed of His residue pairs chelate borderline transition metal ions such as Ni(II). In addition, there are many sulfur-containing donor groups that can coordinate soft metal ions such as Hg(II). Cd(II) may bind to both types of the above listed donor groups. No significant change was observed in the circular dichroism spectra of TEM-1 β-lactamase on increasing the metal ion content of the samples, with the exception of Hg(II) inducing a small change in the secondary structure of the protein. A weak nonspecific binding of Hg(II) was proven by mass spectrometry and 119mHg perturbed angular correlation spectroscopy. The hydrolytic process of ampicillin catalyzed by TEM-1 β-lactamase was described by the kinetic analysis of the set of full catalytic progress curves, where the slow, yet observable conversion of the primary reaction product into a second one, identified as ampilloic acid by mass spectrometry, needed also to be considered in the applied model. Ni(II) and Cd(II) slightly promoted the catalytic activity of the enzyme while Hg(II) exerted a noticeable inhibitory effect. Hg(II) and Ni(II), applied at 10 μM concentration, inhibited the growth of E. coli BL21(DE3) in M9 minimal medium in the absence of ampicillin, but addition of the antibiotic could neutralize this toxic effect by complexing the metal ions.

KW - Hg perturbed angular correlation spectroscopy of γ-rays

KW - circular dichroism

KW - mass spectrometry

KW - reaction kinetics

KW - TEM-1 β-lactamase

KW - toxic metal binding

U2 - 10.1002/pro.4809

DO - 10.1002/pro.4809

M3 - Journal article

C2 - 37853808

AN - SCOPUS:85177553242

VL - 32

JO - Protein Science

JF - Protein Science

SN - 0961-8368

IS - 12

M1 - e4809

ER -