Nucleotide-binding properties of kinase-deficient epidermal-growth-factor-receptor mutants.
Nucleotide-binding properties of kinase-deficient epidermal-growth-factor-receptor mutants.
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激酶缺陷型表皮生长因子受体突变体的核苷酸结合特性。
DOI：
10.1042/bj3300353
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发表时间：
1998
期刊：
影响因子：
--
通讯作者：
Koland,JG
中科院分区：
文献类型：
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作者：
Cheng,K;Koland,JG
采用核苷酸类似物2 <$（3 <$）-O-（2，4，6-trinitrophenyl）adenosine 5 α-triphosphate.测定了野生型和两种突变蛋白的PTK结构域与ATP和Mn·ATP复合物的结合亲和力。令人惊讶的是，在EGF受体PTK结构域的核苷酸结合位点中高度保守的Lys-721残基的突变并没有消除ATP和Mn·ATP结合，尽管对Mn·ATP复合物的结合亲和力显著降低。靶向高度保守的Asp-813残基的第二个激酶失活突变对EGF受体PTK结构域的核苷酸结合特性几乎没有影响。这些结果表明，这两个激酶失活氨基酸取代的主要作用不是阻断核苷酸结合，而是抑制磷酸转移反应。
The nucleotide-binding properties of wild-type epidermal- growth-factor (EGF)-receptor protein tyrosine kinase (PTK) and EGF-receptor mutants with site-specific amino acid substitutions known to attenuate protein kinase activity were analysed by a fluorescence competition assay employing the nucleotide analogue 2ʹ(3ʹ)-O-(2,4,6-trinitrophenyl)adenosine 5ʹ-triphosphate.Binding affinities for ATP and Mn·ATP complex were determined for the PTK domains of the wild-type and two mutant proteins. Surprisingly, mutation of the highly conserved Lys-721 residue in the nucleotide-binding site of the EGF- receptor PTK domain did not abolish ATP and Mn·ATP binding, although the binding affinity for the Mn·ATP complex was significantly reduced. A second kinase-inactivating mutation that targeted the highly conserved Asp-813 residue had little effect on the nucleotide-binding properties of the EGF-receptor PTK domain. These results indicated that the principle effect of these two kinase-inactivating amino acid substitutions is not to block nucleotide binding, but is instead an inhibition of the phospho-transfer reaction.