Del Prete Sonia, Vullo Daniela, di Fonzo Pietro, Carginale Vincenzo, Supuran Claudiu T, Capasso Clemente
Istituto di Bioscienze e Biorisorse, CNR, Via Pietro Castellino 111, 80131 Napoli, Italy; Università degliStudi di Firenze, Dipartimento Di Chimica, Laboratorio di Chimica Bioinorganica, Polo Scientifico, Via della Lastruccia 3, 50019 Sesto Fiorentino, Florence, Italy.
Università degliStudi di Firenze, Dipartimento Di Chimica, Laboratorio di Chimica Bioinorganica, Polo Scientifico, Via della Lastruccia 3, 50019 Sesto Fiorentino, Florence, Italy.
Bioorg Med Chem. 2017 Mar 15;25(6):2010-2015. doi: 10.1016/j.bmc.2017.02.032. Epub 2017 Feb 16.
We report the cloning, purification and characterization of BpsβCA, a β-class carbonic anhydrase (CA, EC 4.2.1.1) from the pathogenic bacterium Burkholderia pseudomallei, the etiological agent of melioidosis, and compare its activity and inhibition with those of the γ-CA from the same organism, BpsγCA, recently investigated by our groups. BpsβCA showed a significant catalytic activity for the physiologic, CO hydration reaction, with the following kinetic parameters, k of 1.6×10s and k/K of 3.4×10M×s. The inhibition of BpsβCA with a group of anions and small molecules was also investigated. The best inhibitors were sulfamide, sulfamic acid and phenylarsonic acid, which showed Ks in the range of 83-92µM, whereas phenylboronic acid, fluoride, cyanide, azide, bisulfite, tetraborate, perrhenate, perruthenate, peroxydisulfate, perchlorate, tetrafluoroborate, fluorosulfonate and hexafluorophosphate showed Ks>100mM. Other inhibitors of this new enzyme were bicarbonate, trithiocarbonate, some complex inorganic anions and N,N-diethyldithiocarbamate, which had inhibition constants of 0.32-8.6mM. As little is known of the life cycle and virulence of this bacterium, this type of study may bring information of interest for the development of novel strategies to fight bacterial infection and drug resistance to commonly used antibiotics.
我们报道了从类鼻疽病原菌——致病性细菌伯克霍尔德菌（Burkholderia pseudomallei）中克隆、纯化并鉴定了一种β类碳酸酐酶（CA，EC 4.2.1.1），即BpsβCA，并将其活性和抑制情况与我们团队最近研究的同一生物体中的γ-CA（BpsγCA）进行了比较。BpsβCA对生理性的CO水合反应表现出显著的催化活性，其动力学参数如下：k为1.6×10⁵ s⁻¹，k/Km为3.4×10⁷ M⁻¹×s⁻¹。我们还研究了一组阴离子和小分子对BpsβCA的抑制作用。最佳抑制剂是氨磺酰胺、氨基磺酸和苯胂酸，其Ks在83 - 92μM范围内，而苯硼酸、氟化物、氰化物、叠氮化物、亚硫酸氢盐、四硼酸盐、高铼酸盐、高钌酸盐、过二硫酸盐、高氯酸盐、四氟硼酸盐、氟磺酸盐和六氟磷酸盐的Ks>100 mM。这种新酶的其他抑制剂包括碳酸氢盐、三硫代碳酸盐、一些复杂的无机阴离子和N,N - 二乙基二硫代氨基甲酸盐，其抑制常数为0.32 - 8.6 mM。由于对这种细菌的生命周期和毒力了解甚少，这类研究可能会为开发对抗细菌感染和常用抗生素耐药性的新策略带来有价值的信息。
