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    • 6. 发明授权
    • Metallo-endopeptidases
    • 金属内肽酶
    • US06518054B1
    • 2003-02-11
    • US09381982
    • 1999-12-07
    • Lambertus Van den BurgOene Robert VeltmanGerard Venema
    • Lambertus Van den BurgOene Robert VeltmanGerard Venema
    • C12N900
    • A23J3/34C11D3/386C11D3/38663C12N9/54C12P21/06C14C1/065
    • The present invention provides genes encoding variants of metallo-endopeptidases that have been engineered to be resistant to prolonged boiling while having maintained their enzymatic performance at much lower temperatures. In addition, thermal stability of the metallo-endopeptidases is highly dependent on calcium at concentrations in the mM range. The invention further provides active metallo-endopeptidases variants whose stability depending on calcium concentration can be changed so as to provide metallo-endopeptidases that are calcium dependent or independent. The invention also provides genes that encode boiling-resistant metallo-endopetidases whose stability depending on calcium concentration can be changed. The invention also provides vectors and cells comprising these genes and proteases produced through these genes, vectors and/or cells. In particular variants with the above described properties are provided of thermolysin-like proteases such as produced by Bacillus stearothermophilus (TLP-ste) and Bacillus thermoproteolyticus (thermolysine). Boiling-resistant and calcium independent or dependent metallo-endopeptidases can be applied in several industrial processes, for instance in the preparation of the artificial sweetener aspartame, but also in the leather industry, in breweries and in the production of protein hydrolysates.
    • 本发明提供编码金属内肽酶变体的基因,其已被工程化以对延长的煮沸有抵抗力,同时保持其在较低温度下的酶性能。 此外,金属内肽酶的热稳定性高度依赖于在mM范围内的浓度下的钙。 本发明还提供活性金属内肽酶变体,其可以改变其依赖于钙浓度的稳定性,以提供钙依赖性或独立性的金属内肽酶。 本发明还提供编码耐沸腾金属内肽酶的基因,其可以改变其依赖于钙浓度的稳定性。 本发明还提供了通过这些基因,载体和/或细胞产生的包含这些基因和蛋白酶的载体和细胞。 特别是具有上述性质的变体提供了嗜热脂肪芽孢杆菌(Bacillus stearothermophilus)(TLP-ste)和热嗜热芽孢杆菌(Thermolycolyticus)产生的嗜热菌蛋白酶样蛋白酶。 抗沸腾和钙依赖性或依赖性金属内肽酶可以应用于几个工业过程中,例如制备人造甜味剂阿斯巴甜,还可用于皮革工业,酿酒厂和生产蛋白质水解产物。