Lehtonen Jonna, Shen Bairong, Vihinen Mauno, Casini Angela, Scozzafava Andrea, Supuran Claudiu T, Parkkila Anna-Kaisa, Saarnio Juha, Kivelä Antti J, Waheed Abdul, Sly William S, Parkkila Seppo
Institute of Medical Technology, University of Tampere, Tampere, Finland.
J Biol Chem. 2004 Jan 23;279(4):2719-27. doi: 10.1074/jbc.M308984200. Epub 2003 Nov 4.
The carbonic anhydrase (CA) gene family has been reported to consist of at least 11 enzymatically active members and a few inactive homologous proteins. Recent analyses of human and mouse databases provided evidence that human and mouse genomes contain genes for still another novel CA isozyme hereby named CA XIII. In the present study, we modeled the structure of human CA XIII. This model revealed a globular molecule with high structural similarity to cytosolic isozymes, CA I, II, and III. Recombinant mouse CA XIII showed catalytic activity similar to those of mitochondrial CA V and cytosolic CA I, with k(cat)/K(m) of 4.3 x 10(7) m(-1) s(-1), and k(cat) of 8.3 x 10(4) s(-1). It is very susceptible to inhibition by sulfonamide and anionic inhibitors, with inhibition constants of 17 nm for acetazolamide, a clinically used sulfonamide, and of 0.25 microm, for cyanate, respectively. Using panels of cDNAs we evaluated human and mouse CA13 gene expression in a number of different tissues. In human tissues, positive signals were identified in the thymus, small intestine, spleen, prostate, ovary, colon, and testis. In mouse, positive tissues included the spleen, lung, kidney, heart, brain, skeletal muscle, and testis. We also investigated the cellular and subcellular localization of CA XIII in human and mouse tissues using an antibody raised against a polypeptide of 14 amino acids common for both human and mouse orthologues. Immunohistochemical staining showed a unique and widespread distribution pattern for CA XIII compared with the other cytosolic CA isozymes. In conclusion, the predicted amino acid sequence, structural model, distribution, and activity data suggest that CA XIII represents a novel enzyme, which may play important physiological roles in several organs.
据报道，碳酸酐酶（CA）基因家族至少由11个具有酶活性的成员和一些无活性的同源蛋白组成。最近对人类和小鼠数据库的分析表明，人类和小鼠基因组中还含有另一种新型CA同工酶的基因，命名为CA XIII。在本研究中，我们构建了人类CA XIII的结构模型。该模型显示，其为球状分子，与胞质同工酶CA I、II和III具有高度的结构相似性。重组小鼠CA XIII表现出与线粒体CA V和胞质CA I相似的催化活性，催化常数（k(cat)/K(m)）为4.3×10(7) m(-1) s(-1)，催化速率常数（k(cat)）为8.3×10(4) s(-1)。它对磺胺类和阴离子抑制剂非常敏感，临床使用的磺胺类药物乙酰唑胺的抑制常数为17 nm，氰酸盐的抑制常数为0.25 μm。我们使用一系列cDNA评估了人类和小鼠CA13基因在多种不同组织中的表达。在人类组织中，在胸腺、小肠、脾脏、前列腺、卵巢、结肠和睾丸中检测到阳性信号。在小鼠中，阳性组织包括脾脏、肺、肾、心脏、脑、骨骼肌和睾丸。我们还使用针对人类和小鼠直系同源物共有的14个氨基酸多肽产生的抗体，研究了人类和小鼠组织中CA XIII的细胞和亚细胞定位。免疫组织化学染色显示，与其他胞质CA同工酶相比，CA XIII具有独特且广泛的分布模式。总之，预测的氨基酸序列、结构模型、分布和活性数据表明，CA XIII代表一种新型酶，可能在多个器官中发挥重要的生理作用。
