Functional significance of lysine 1423 of neurofibromin and characterization of a second site suppressor which rescues mutations at this residue and suppresses RAS2Val-19-activated phenotypes.
Functional significance of lysine 1423 of neurofibromin and characterization of a second site suppressor which rescues mutations at this residue and suppresses RAS2Val-19-activated phenotypes.
复制标题
神经纤维蛋白 1423 赖氨酸的功能意义以及第二位点抑制因子的表征，该抑制因子可挽救该残基的突变并抑制 RAS2Val-19 激活的表型。
DOI：
10.1128/mcb.14.1.815-821.1994
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发表时间：
1994
影响因子：
5.3
通讯作者：
Tamanoi,F
中科院分区：
文献类型：
--
作者：
Poullet,P;Lin,B;Esson,K;Tamanoi,F
神经纤维蛋白（神经纤维瘤病I型基因产物[NF 1]）的赖氨酸1423在NF 1的功能中起着至关重要的作用。在神经纤维瘤病患者和癌症患者的样品中检测到这种赖氨酸的突变。为了进一步了解这个残基的重要性，我们将其突变为所有可能的氨基酸。使用酵母互补的功能测定已经揭示赖氨酸是产生功能性NF 1的唯一氨基酸。不同突变蛋白的定量分析表明，它们的GTP酶激活蛋白（GAP）活性急剧下降，作为其Ras亲和力下降的结果。在GAP相关结构域内的其他保守残基的情况下，没有观察到对特定残基的这种需要。我们还报告，另一个残基，苯丙氨酸1434，在NF 1功能中起着重要作用。这首先由以下发现表明：由于赖氨酸1423改变为其他氨基酸而导致的缺陷性NF 1可以通过残基1434处的第二位点基因内突变来挽救。该突变部分恢复了赖氨酸突变体中的GAP活性。当将苯丙氨酸1434突变为丝氨酸引入野生型NF 1蛋白时，所得蛋白获得了抑制RAS 2 Val-19细胞活化表型的能力。然而，这种抑制不涉及Ras相互作用，因为苯丙氨酸突变体不刺激RAS 2 Val-19蛋白的内在GT3活性，并且对Ras蛋白的亲和力不增加。
Lysine 1423 of neurofibromin (neurofibromatosis type I gene product [NF1]) plays a crucial role in the function of NF1. Mutations of this lysine were detected in samples from a neurofibromatosis patient as well as from cancer patients. To further understand the significance of this residue, we have mutated it to all possible amino acids. Functional assays using yeastiracomplementation have revealed that lysine is the only amino acid that produced functional NF1. Quantitative analyses of different mutant proteins have suggested that their GTPase-activating protein (GAP) activity is drastically reduced as a result of a decrease in their Ras affinity. Such a requirement for a specific residue is not observed in the case of other conserved residues within the GAP-related domain. We also report that another residue, phenylalanine 1434, plays an important role in NF1 function. This was first indicated by the finding that defective NF1s due to an alteration of lysine 1423 to other amino acids can be rescued by a second site intragenic mutation at residue 1434. The mutation partially restored GAP activity in the lysine mutant. When the mutation phenylalanine 1434 to serine was introduced into a wild-type NF1 protein, the resulting protein acquired the ability to suppress activated phenotypes ofRAS2Val-19cells. This suppression, however, does not involve Ras interaction, since the phenylalanine mutant does not stimulate the intrinsic GTPase activity of RAS2Val-19protein and does not have an increased affinity for Ras proteins.