BIOCHEMICAL GENETICS OF CARBONIC ANHYDRASE DEFICIENCIES

这项研究的广泛目标是继续研究遗传性 缺乏人类碳酸酐酶（CA），以确定 该基因家族的单个成员的生理作用， 破译它们的遗传和生理相互关系， 了解他们对健康和疾病的重要性。 本项目期间有五个具体目标： 1)完整的研究定义的生化和分子遗传学 碳酸酐酶II缺乏症。(CA第二，不足是基础。 新分类的先天性缺陷性肾小管性石骨症 酸中毒和脑钙化。） 2)完成碳酸酐酶的生化和分子遗传学研究 四. (CA IV是肾和肺中的膜锚定CA， 毛细血管内皮细胞的管腔表面）。 3)通过靶向突变建立CA IV缺乏症小鼠模型 在ES细胞中。 4)定义CA V的生物化学和分子遗传学。（CA V是 （在肝脏和肌肉中的核编码的线粒体CA）。 5)通过靶向诱变产生小鼠CA V缺陷模型， ES细胞。 碳酸酐酶被纯化、表征、经受 微测序，并用于制备多克隆抗体的代谢 标记和生物合成研究。 cDNA和基因组克隆是 分离以确定基因组组织，内含子序列， 周围的内含子/外显子边界，染色体定位，以及 突变的本质改变了碳链的表达， 脱水酶基因 ES细胞中的靶向诱变将被利用， 生产动物模型的选择CA缺陷。

The broad goal of this research is to continue studies of inherited deficiencies of human carbonic anhydrases (CAs) in order to define the physiological roles of individual members of this gene family, to decipher their genetic and physiologic interrelationships, and to understand their individual importance in health and disease. For this project period there are five specific aims: 1) Complete studies defining the biochemical and molecular genetics of carbonic anhydrase II deficiency. (CA II deficiency is the basis of the newly classified inborn error producing osteopetrosis, renal tubular acidosis, and brain calcification.) 2) Complete the biochemical and molecular genetics of carbonic anhydrase IV. (CA IV is the membrane anchored CA in kidney and lung and on the luminal surface of capillary endothelial cells). 3) Produce a murine model for CA IV deficiency by targeted mutagenesis in ES cells. 4) Define the biochemical and molecular genetics of CA V. (CA V is the nuclear-encoded, mitochondrial CA in liver and muscle.) 5) Produce a murine model of CA V deficiency by targeted mutagenesis in ES cells. Carbonic anhydrases are purified, characterized, subjected to microsequencing, and used to prepare polyclonal antibodies for metabolic labeling and biosynthetic studies. The cDNA and genomic clones are isolated to determine the genomic organization, the intronic sequences surrounding intron/exon boundaries, the chromosomal localization, and the nature of the mutations underlying altered expression of the carbonic anhydrase genes. Targeted mutagenesis in ES cells will be exploited to produce animal models of selected CA deficiencies.