167 research outputs found
Lu et al-An Active Site Loop Toggles Between Conformations to Control Antibiotic Hydrolysis and Inhibition Potency for CTX-M beta-lactamase Drug-Resistance Enzymes
No full textMolecular dynamics simulations trajectories for publication, "An Active Site Loop Toggles Between Conformations to Control Antibiotic Hydrolysis and Inhibition Potency for CTX-M beta-lactamase Drug-Resistance Enzymes"
Shuo Lu1, Liya Hu2, Hanfeng Lin1, Allison Judge2, Paola Rivera1, Murugesan Palaniappan3, Banumathi Sankaran4, Jin Wang1, B.V. Venkataram Prasad2 and Timothy Palzkill1,2*
Department of Pharmacology and Chemical Biology1, Department of Biochemistry and Molecular Biology2, Center for Drug Discovery, Department of Pathology & Immunology3, Baylor College of Medicine, Houston, TX 77030
Department of Molecular Biophysics and Integrated Bioimaging, Berkeley Center for Structural Biology, Lawrence Berkeley National Laboratory, Berkeley, California, USA4
*Corresponding author: [email protected]</p
800 QM/MM minimum energy pathway conformations for the deacylation reactions of KPC-2/Imipenem
No full textThe datasets comprise 800 coordinate files (in the CHARMM psf/cor format) for the QM/MM minimum energy pathways of the deacylation reactions between a Class A beta-lactamases (KPC wild type (KPC-WT) and KPC Phe72Tyr mutant (KPC-F72Y)) and the imipenem antibiotic molecues.
All pathway conformations were optimized at DFTB3/3OB-f/CHARMM36 level with 36 replicas.
All single point calculations and charge population analysis were done at B3LYP-D3/6-31+G(d,p)/CHARMM36 level.
0.paths_kpc_f72y_ipm_d1.tar.gz: 200 pathways conformations for KPC-F72Y/IPM-Delta1 deacylation reactions
0.paths_kpc_f72y_ipm_d2.tar.gz: 200 pathways conformations for KPC-F72Y/IPM-Delta2 deacylation reactions
0.paths_kpc_wt_ipm_d1.tar.gz: 200 pathways conformations for KPC-WT/IPM-Delta1 deacylation reactions
0.paths_kpc_wt_ipm_d2.tar.gz: 200 pathways conformations for KPC-WT/IPM-Delta2 deacylation reactions
1.chrgs.tar.gz: The CHELPG charges of the QM region of all replica conformations along all KPC-2/IPM pathways.
2.enes.tar.gz: The single point replica energies along all KPC-2/IPM pathways.
3.datasets.tar.gz: The 16 features dataset used in this project and SHAP value for every feature.
4.code.tar.gz: Code for figures plotting, models training with four different ML model and hydrogen bonds searching using MDAnalysis.
5.dftb_ti.tar.gz: Structure and logs files for free energy calculation for KPC-WT/IPM-Delta1 and KPC-WT/IPM-Delta2
Structural and Mechanistic Basis for Extended-Spectrum Drug-Resistance Mutations in Altering the Specificity of TEM, CTX-M, and KPC β-lactamases
No full textThe most common mechanism of resistance to β-lactam antibiotics in Gram-negative bacteria is the production of β-lactamases that hydrolyze the drugs. Class A β-lactamases are serine active-site hydrolases that include the common TEM, CTX-M, and KPC enzymes. The TEM enzymes readily hydrolyze penicillins and older cephalosporins. Oxyimino-cephalosporins, such as cefotaxime and ceftazidime, however, are poor substrates for TEM-1 and were introduced, in part, to circumvent β-lactamase-mediated resistance. Nevertheless, the use of these antibiotics has lead to evolution of numerous variants of TEM with mutations that significantly increase the hydrolysis of the newer cephalosporins. The CTX-M enzymes emerged in the late 1980s and hydrolyze penicillins and older cephalosporins and derive their name from the ability to also hydrolyze cefotaxime. The CTX-M enzymes, however, do not efficiently hydrolyze ceftazidime. Variants of CTX-M enzymes, however, have evolved that exhibit increased hydrolysis of ceftazidime. Finally, the KPC enzyme emerged in the 1990s and is characterized by its broad specificity that includes penicillins, most cephalosporins, and carbapenems. The KPC enzyme, however, does not efficiently hydrolyze ceftazidime. As with the TEM and CTX-M enzymes, variants have recently evolved that extend the spectrum of KPC β-lactamase to include ceftazidime. This review discusses the structural and mechanistic basis for the expanded substrate specificity of each of these enzymes that result from natural mutations that confer oxyimino-cephalosporin resistance. For the TEM enzyme, extended-spectrum mutations act by establishing new interactions with the cephalosporin. These mutations increase the conformational heterogeneity of the active site to create sub-states that better accommodate the larger drugs. The mutations expanding the spectrum of CTX-M enzymes also affect the flexibility and conformation of the active site to accommodate ceftazidime. Although structural data are limited, extended-spectrum mutations in KPC may act by mediating new, direct interactions with substrate and/or altering conformations of the active site. In many cases, mutations that expand the substrate profile of these enzymes simultaneously decrease the thermodynamic stability. This leads to the emergence of additional global suppressor mutations that help correct the stability defects leading to increased protein expression and increased antibiotic resistance
Structural and Mechanistic Basis for Extended-Spectrum Drug-Resistance Mutations in Altering the Specificity of TEM, CTX-M, and KPC β-lactamases
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Una prueba para la detección rápida del efecto de inóculo de la cefazolina en Staphylococcus aureus sensible a la meticilina
No full textEl efecto inóculo de la cefazolina (CzIE) se ha asociado a fracasos terapéuticos y mortalidad en infecciones invasivas por Staphylococcus aureus sensible a la meticilina (MSSA). Actualmente no se dispone de una prueba diagnóstica para detectar el CzIE. Desarrollamos una prueba colorimétrica CzIE rápida (∼3 h) para detectar la actividad estafilocócica-β-lactamasa (BlaZ) en sobrenadantes tras la inducción con ampicilina. La prueba se validó utilizando 689 aislados de MSSA del torrente sanguíneo recuperados de Latinoamérica y Estados Unidos. Se utilizó como patrón de referencia la determinación de la CMI de cefazolina con un inóculo elevado (107 UFC/ml) (punto de corte ≥16 μg/ml). Todos los aislados se sometieron a secuenciación del genoma. Un total de 257 (37,3%) de los aislados de MSSA presentaron la CzIE por el método del estándar de referencia. La sensibilidad y especificidad globales de la prueba colorimétrica fueron del 82,5% y el 88,9%, respectivamente. La sensibilidad en los aislados de MSSA que albergaban BlaZ de tipo A (la enzima más eficaz contra la cefazolina) fue del 92,7%, con una especificidad del 87,8%. El rendimiento de la prueba fue inferior frente a las enzimas de tipo B y C (sensibilidades del 53,3% y el 72,3%, respectivamente). Cuando el valor de referencia se fijó en ≥32 μg/ml, la sensibilidad para los aislados portadores de enzimas de tipo A fue del 98,2%. La especificidad fue del 100% para el MSSA carente de blaZ. El valor predictivo negativo global osciló entre el 81,4% y el 95,6% en los países latinoamericanos utilizando las tasas de prevalencia publicadas de la CzIE. Los aislados de MSSA de los Estados Unidos eran genéticamente diversos, sin diferencias genómicas distintivas con respecto al MSSA latinoamericano, distribuidos entre 18 tipos de secuencia. Una nueva prueba puede identificar fácilmente la mayoría de los aislados de MSSA que presentan la CzIE, en particular los portadores del tipo A BlaZ. A diferencia de la determinación de la CMI utilizando un inóculo elevado, la prueba rápida es barata, factible y fácil de realizar. Tras unos pequeños pasos de validación, podría incorporarse al flujo de trabajo rutinario del laboratorio clínico.The cefazolin inoculum effect (CzIE) has been associated with therapeutic failures and mortality in invasive methicillin-susceptible Staphylococcus aureus (MSSA) infections. A diagnostic test to detect the CzIE is not currently available. We developed a rapid (∼3 h) CzIE colorimetric test to detect staphylococcal-β-lactamase (BlaZ) activity in supernatants after ampicillin induction. The test was validated using 689 bloodstream MSSA isolates recovered from Latin America and the United States. The cefazolin MIC determination at a high inoculum (107 CFU/ml) was used as a reference standard (cutoff ≥16 μg/ml). All isolates underwent genome sequencing. A total of 257 (37.3%) of MSSA isolates exhibited the CzIE by the reference standard method. The overall sensitivity and specificity of the colorimetric test was 82.5% and 88.9%, respectively. Sensitivity in MSSA isolates harboring type A BlaZ (the most efficient enzyme against cefazolin) was 92.7% with a specificity of 87.8%. The performance of the test was lower against type B and C enzymes (sensitivities of 53.3% and 72.3%, respectively). When the reference value was set to ≥32 μg/ml, the sensitivity for isolates carrying type A enzymes was 98.2%. Specificity was 100% for MSSA lacking blaZ. The overall negative predictive value ranged from 81.4% to 95.6% in Latin American countries using published prevalence rates of the CzIE. MSSA isolates from the United States were genetically diverse, with no distinguishing genomic differences from Latin American MSSA, distributed among 18 sequence types. A novel test can readily identify most MSSA isolates exhibiting the CzIE, particularly those carrying type A BlaZ. In contrast to the MIC determination using high inoculum, the rapid test is inexpensive, feasible, and easy to perform. After minor validation steps, it could be incorporated into the routine clinical laboratory workflow
Dissecting the Protein-Protein Interface between β-Lactamase Inhibitory Protein and Class A β-Lactamases
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