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    • 16. 发明申请
    • NOVEL THERMOLYSIN-LIKE PROTEASE AND USE THEREOF
    • 新型热病菌样蛋白及其用途
    • WO1997021804A1
    • 1997-06-19
    • PCT/JP1996003617
    • 1996-12-11
    • SAGAMI CHEMICAL RESEARCH CENTERHOLLAND SWEETENER COMPANY V.O.F.HANZAWA, SatoshiOHE, SeigoMIYAKE, ToshioKIDOKORO, Shun-ichiENDO, KimikoWADA, Akiyoshi
    • SAGAMI CHEMICAL RESEARCH CENTERHOLLAND SWEETENER COMPANY V.O.F.
    • C12N09/54
    • C07K5/0613C12N9/54C12Y304/24027
    • A novel thermolysin-like protease having an improved stability and a method wherein this protease is used in a coupling reaction with phenylalanine methyl ester so as to elevate the efficiency of the reaction. The novel thermolysin-like protease is one which is to be used in the coupling of N-benzyloxycarbonylaspartic acid with phenylalanine methyl ester in a water soluble medium in a process for synthesizing benzyloxycarbonyl- alpha -L-aspartyl-L-phenylalanine methyl ester via the enzymatic reaction and characterized in that the volume of a void in electron density existing in the thermolysin having the amino acid sequence of SEQ ID NO:1 in the Sequence Listing and surrounded at least by Leu at the 144-position, Thr at the 149-position, Ala at the 240-position, Ala at the 270-position and Ala at the 288-position has been at least partly made up for by substituting at least one of these amino acid residues by amino acid residue(s) having larger hydrophobicity and bulkiness, or by affecting the amino acid residues surrounding the void in electron density so as to reduce the volume of the void, and a method wherein the protease is used in the coupling reaction of N-benzyloxycarbonylaspartic acid with phenylalanine methyl ester.
    • 具有改进的稳定性的新型嗜热菌蛋白酶样蛋白酶和其中所述蛋白酶用于与苯丙氨酸甲酯的偶联反应中以提高反应效率的方法。 新型嗜热溶蛋白酶样蛋白酶是一种用于在苄基氧羰基-α-天冬氨酰基-L-苯丙氨酸甲酯合成方法中通过水溶性介质偶联N-苄氧基羰基天冬氨酸与苯丙氨酸甲酯的蛋白酶 酶反应,其特征在于存在于序列表中具有SEQ ID NO:1的氨基酸序列的嗜热溶菌素中的电子密度的空隙的体积,至少被144位的Leu,在149位的Thr, 位置,240位的Ala,270位的Ala和288位的Ala已经至少部分地由这些氨基酸残基中的至少一个用疏水性较大的氨基酸残基代替 或通过以电子密度影响空隙周围的氨基酸残基以减小空隙体积的方法,以及其中所述蛋白酶用于N-苄氧基羰基天冬氨酸的偶联反应的方法 与苯丙氨酸甲酯。