公开(公告)号：KR100732604B1

公开(公告)日：2007-06-27

申请号：KR1020060013726

申请日：2006-02-13

Applicant: 한국과학기술연구원

Inventor： 김은경 ,  양은경 ,  황광연 ,  이원호 ,  한기철

IPC: C12N15/54 ,  C12N9/12

Abstract: A modified 5-AMP(adenine monophosphate)-activated protein kinase(AMPK), a method for preparing the same protein kinase, and use thereof are provided to confer the water-soluble and crystalline structure which is more easily analyzed in the three-dimensional structure analysis on the protein kinase, so that the modified protein kinase acts as a target protein of obesity treatment. The method for preparing the modified water-soluble and crystalline 5-AMP-activated protein kinase comprises the steps of: (i) preparing water-soluble modified AMPK protein wherein a preserved protein kinase active domain of AMPK alpha subunit having the amino acid sequence of SEQ ID NO:1 is contained, and one polypeptide molecule selected from 1-12 amino acid sequences from the N-terminal region is deleted, one polypeptide molecule selected from 250-280 amino acid sequences from the C-terminal region is deleted, or both the polypeptide molecules are deleted; and (ii) crystallizing the modified AMPK protein by performing a hanging-drop vapour diffusion method using water tank solution containing a precipitating agent selected from ammonium sulfate and MES(Methyl ester sulfonate) at 16-26 deg.C and pH 6.0-6.5.

Abstract translation: 提供了修饰的5-AMP（腺嘌呤单磷酸）激活的蛋白激酶（AMPK），制备相同蛋白激酶的方法及其用途，以赋予在三维中更容易分析的水溶性和晶体结构 对蛋白激酶进行结构分析，使修饰的蛋白激酶作为肥胖治疗的靶蛋白。 制备修饰的水溶性和结晶5-AMP活化蛋白激酶的方法包括以下步骤：（i）制备水溶性修饰的AMPK蛋白，其中AMPKα亚基的保守蛋白激酶活性结构域具有氨基酸序列 含有SEQ ID NO：1，并且选自N末端区域的1-12个氨基酸序列中的一个多肽分子缺失，选自C末端区域的250-280个氨基酸序列中的一个多肽分子缺失，或者 多肽分子都被删除; （ii）通过使用含有选自硫酸铵和MES（甲基酯磺酸盐）的沉淀剂的水罐溶液在16-26℃和pH 6.0-6.5下进行悬滴蒸气扩散方法使修饰的AMPK蛋白质结晶。