SITE-DIRECTED MUTAGENIC REPLACEMENT OF GLU-461 WITH GLN IN BETA-GALACTOSIDASE (ESCHERICHIA-COLI) - EVIDENCE THAT GLU-461 IS IMPORTANT FOR ACTIVITY
SITE-DIRECTED MUTAGENIC REPLACEMENT OF GLU-461 WITH GLN IN BETA-GALACTOSIDASE (ESCHERICHIA-COLI) - EVIDENCE THAT GLU-461 IS IMPORTANT FOR ACTIVITY
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DOI：
10.1016/s0006-291x(88)81222-1
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发表时间：
1988-05-31
影响因子：
3.1
通讯作者：
HUBER, RE
中科院分区：
文献类型：
--
作者：
BADER, DE;RING, M;HUBER, RE
谷氨酸461的。-半乳糖苷酶（大肠杆菌）通过定点诱变被谷氨酰胺取代。纯化Q461- β的动力学研究。-半乳糖苷酶显示其活性低于野生型（以ONPG为底物）的0.4%，证实了顺序研究表明葡萄糖-461上的负电荷对活性很重要。Km值没有增加，表明底物的结合没有因为这种变化而降低。热变性研究表明Q461- β。-半乳糖苷酶比野生型酶更容易热变性。
Glutamic acid 461 of .beta.-galactosidase (E. coli) was replaced by gln using site-directed mutagenesis. Kinetic studies on the purified Q461-.beta.-galactosidase showed that it had < 0.4% of the wild-type activity (with ONPG as substrate), confirming order studies which have suggested that the negative charge on glu-461 is important for activity. The Km values did not increase, indicating that binding of the substrate was not decreased by this change. Thermal denaturation studies showed Q461-.beta.-galactosidase to be somewhat more susceptible to heat denaturation than the wild-type enzyme.