Nishimori Isao, Minakuchi Tomoko, Onishi Saburo, Vullo Daniela, Cecchi Alessandro, Scozzafava Andrea, Supuran Claudiu T
Department of Gastroenterology and Hepatology, Kochi Medical School, Nankoku, Kochi 783-8505, Japan.
Bioorg Med Chem. 2007 Dec 1;15(23):7229-36. doi: 10.1016/j.bmc.2007.08.037. Epub 2007 Aug 25.
The cytosolic human carbonic anhydrase (hCA, EC 4.2.1.1) isozyme III (hCA III) has been cloned and purified by the GST-fusion protein method. Recombinant pure hCA III had the following kinetic parameters for the CO(2) hydration reaction at 20 degrees C and pH 7.5: k(cat) of 1.3 x 10(4) s(-1) and k(cat)/K(M) of 2.5 x 10(5) M(-1) s(-1), being a slower catalyst for the physiological reaction as compared to the genetically related cytosolic isoforms hCA I and II. An inhibition study with a library of sulfonamides and one sulfamate, some which are clinically used compounds, is reported. hCA III is less prone to be inhibited by these compounds as compared to hCA I and II for which many low nanomolar inhibitors were detected earlier. The best hCA III inhibitors were prontosil, sulpiride, indisulam, benzolamide, aminobenzolamide, and 4-amino-6-chloro-benzene-1,3-disulfonamide which showed K(I)s in the range of 2.3-18.1 microM. Clinically used compounds such as acetazolamide, methazolamide, ethoxzolamide, dorzolamide, brinzolamide, topiramate, zonisamide, celecoxib, and valdecoxib were less effective hCA III inhibitors, with affinities in the range of 154-2200 microM. This is the first study in which low micromolar hCA III inhibitors are reported.
胞质型人碳酸酐酶（hCA，EC 4.2.1.1）同工酶III（hCA III）已通过谷胱甘肽S-转移酶（GST）融合蛋白方法进行克隆和纯化。重组纯hCA III在20℃和pH 7.5条件下对CO₂水合反应具有以下动力学参数：催化常数（kcat）为1.3×10⁴ s⁻¹，催化效率（kcat/KM）为2.5×10⁵ M⁻¹ s⁻¹，与遗传相关的胞质同工型hCA I和II相比，它是生理反应的较慢催化剂。本文报道了对一系列磺胺类化合物和一种氨基磺酸酯的抑制研究，其中一些是临床使用的化合物。与hCA I和II相比，hCA III较不易被这些化合物抑制，此前已检测到许多低纳摩尔抑制剂可抑制hCA I和II。hCA III的最佳抑制剂是百浪多息、舒必利、茚地那韦、苯并酰胺、氨基苯并酰胺和4-氨基-6-氯苯-1,3-二磺酰胺，其抑制常数（KI）在2.3 - 18.1 μM范围内。临床使用的化合物如乙酰唑胺、甲醋唑胺、乙氧唑胺、多佐胺、布林佐胺、托吡酯、唑尼沙胺、塞来昔布和伐地昔布是效果较差的hCA III抑制剂，亲和力在154 - 2200 μM范围内。这是首次报道低微摩尔浓度hCA III抑制剂的研究。
