Development of novel anti-uricosuric agents based on the genomic strategy.

与大多数其他哺乳动物不同，尿酸盐是高等灵长类动物体内嘌呤降解的主要惰性最终产物，因为肝脏氧化酶尿酸酶基因沉默。肾脏在尿酸盐排出中起主导作用；它排泄每日尿酸盐产量的70%。因此，了解肾尿酸盐的处理机制是很重要的，因为尿酸排泄不足与高尿酸血症的发展有关，从而导致痛风。尿酸盐转运系统存在于近端小管，但由于其双向转运和物种差异，其复杂性较高。最近，我们在人类肾脏中发现了尿酸-阴离子交换因子URAT1(SLC22A12)，发现SLC22A12的缺陷会导致特发性肾脏低尿酸血症。URAT1是影响尿酸盐排泄的尿酸和抗尿酸药物的靶点。尿酸盐转运蛋白的分子鉴定将有助于高尿酸血症新药的开发。我们还发现了…此外，SLC22A12基因缺陷可导致特发性肾功能低尿酸血症(MIM值为220150)，其患者尿酸排泄率高达95±10%(正常为10%)。这些结果表明，URAT1调节血尿酸水平，反之亦然，即要控制血尿酸水平，就需要调节URAT1的转运功能。一例特发性肾功能低尿酸血症患者SLC22A12中新发现的基因突变促使我们考虑URAT1细胞内极端C末端区域对其功能的重要性。URAT1C-末端(1639-1643del)附近5个碱基的缺失会导致移码，末端的7个氨基酸变成8个不同的氨基酸。在非洲爪哇卵母细胞表达系统中，该突变的URAT1转运活性较低。有趣的是，URAT1 C-末端的PDZ结合基序被这种氨基酸序列修饰而消失。在这项研究中，我们使用酵母双杂交方法来研究可能与URAT1相关的调节其运输功能的蛋白质。我们鉴定了含有PDZ结构域的多价蛋白PDZK1是URAT1在人类肾脏中的一个明显的伙伴。此外，我们还发现了PDZK1/URAT1相互作用在HEK293细胞中的功能结果，通过共表达PDZK1，URAT1的转运活性增加了1.4倍。我们推测PDZK1是一种支架蛋白，可能是URAT1功能的生理调节因子。较少

Urate is the major inert end product of purine degradation in higher primates in contrast to most other mammals because of the genetic silencing of hepatic oxidative enzyme uricase. The kidney plays a dominant role in urate elimination ; it excretes 70% of the daily urate production. Therefore, it is important to understand renal urate handling mechanism because the underexcretion of urate has been implicated in the development of hyperuricemia that leads to gout. The urate transport systems exist in the proximal tubule but they are complicated because of their bidirectional transport and the species differences. Recently, we have identified the urate-anion exchanger URAT1 (SLC22A12) in the human kidney and found that defects in SLC22A12 lead to idiopathic renal hypouricemia. URAT1 is targeted by uricosuric and antiuricosuric agents that affect urate excretion. Molecular identification of urate transporting proteins will lead to the new drug development for hyperuricemia.We also found … More that defects in SLC22A12 lead to idiopathic renal hypouricaemia (MIM number, 220150) and patients with such defects show a high fractional urate excretion such as 95 ± 10 % (normally <10%). These results indicate that URAT1 regulates blood urate level and vice versa, that is, to control blood urate levels, the URAT1 transport function should be regulated. A newly found genetic alteration in SLC22A12 from a patient of an idiopathic renal hypouricaemia has prompted us to consider the importance of the URAT1 extreme intracellular C-terminal region for its function. A 5-bp deletion near the URAT1 C-terminal end (1639-1643del) causes frameshift and the seven amino acids in the terminal sequences have changed into eight different amino acids. The URAT1 transport activity of this mutation is low in the Xenopus oocyte expression system. Interestingly, the PDZ binding motif at the C-terminal end of URAT1, which is known to participate in protein-protein interaction, disappears by this amino acid sequence modification.In this study, we use the yeast two-hybrid approach to investigate the putative URAT1-associated proteins that modulate its transport function. We identify the multivalent PDZ domain-containing protein PDZK1 as an apparent partner of URAT1 in the human kidney. Moreover, we show a functional consequence of PDZK1/URAT1 interaction in transfected HEK293 cells, where URAT1 transport activities were increased by 1.4-fold by coexpression of PDZK1. We speculate that PDZK1 is a scaffolding protein that may be a physiological regulator of the function of URAT1. Less