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首页> 外文期刊>Journal of Molecular Biology >Isolation and characterization of mutated mitochondrial GTP : AMP phosphotransferase
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Isolation and characterization of mutated mitochondrial GTP : AMP phosphotransferase

机译：突变线粒体GTP：AMP磷酸转移酶的分离与鉴定

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GTP:AMP phosphotransferase (adenylate kinase isozyme 3, AK3) mutants were obtained by using the ability of AK3 to complement a temperature-sensitive mutation of Escherichia coli adenylate kinase (AKe). Five mutants, P16L, G19S, G91D, G91S, and P93L, had mutation sites located at two loops that are involved in substrate binding of the enzyme. P16L and G19S bearing changes at the first loop showed reduced affinity for both GTP and AMP, the extent of reduction being slightly higher for Gm than AMP. Ln contrast, G91S and P93L having alterations at the second loop had lower affinities for AMP. Only the alterations at the second loop strongly influenced the V-max value of the enzyme. Another mutant, D163N, had a substitution at the site forming a salt bridge in adenylate kinase isozyme 1 (AK1), which influenced the V-max as well as the K-m values for both substrates. The kinetic characteristics of these mutants were comparable to those of the corresponding AK1 or AKe mutants. Furthermore, from the results of mutations T201P and T201A that had alterations in all the kinetic parameters of AK3 and from a comparison with the structure and the kinetic parameters of AKe, we expect that a residue(s) around Thr201 is involved in recognition of the base of nucleoside triphosphate. (C) 1998 Academic Press. [References: 35]
机译：GTP：AMP磷酸转移酶（腺苷酸激酶同工酶3，AK3）突变体是通过利用AK3补充大肠杆菌腺苷酸激酶（AKe）的温度敏感突变而获得的。 P16L，G19S，G91D，G91S和P93L这五个突变体的突变位点位于与酶的底物结合有关的两个环上。在第一个环上的P16L和G19S轴承变化显示出对GTP和AMP的亲和力降低，Gm的降低程度略高于AMP。相比之下，在第二个循环处发生变化的G91S和P93L对AMP的亲和力较低。仅第二个环的改变强烈影响酶的V-max值。另一个突变体D163N在取代腺苷酸激酶同工酶1（AK1）中形成盐桥的位点具有取代基，这影响了两种底物的V-max和K-m值。这些突变体的动力学特性与相应的AK1或AKe突变体相当。此外，根据突变AK2的所有动力学参数均发生变化的T201P和T201A突变的结果，以及与AKe的结构和动力学参数的比较，我们预计Thr201周围的残基参与了对AK3的识别。三磷酸核苷的碱基。 （C）1998年学术出版社。 [参考：35]

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