Zhong J M, Chen-Hwang M C, Hwang Y W
Molecular Biology Department, New York State Institute for Basic Research in Developmental Disabilities, Staten Island 10314, USA.
J Biol Chem. 1995 Apr 28;270(17):10002-7. doi: 10.1074/jbc.270.17.10002.
We examined c-Ha-Ras harboring an aspartate to asparagine substitution at position 119 (mutation D119N). The Asp-119 is part of the conserved NKXD motif shared by members of the regulatory GTPase family. This asparagine residue has been proposed to participate in direct bonding to the guanine ring and to determine the guanine-nucleotide binding specificity. The D119N mutation was found to alter nucleotide specificity of Ha-Ras from guanine to xanthine, an observation that directly supports the essential role of hydrogen bonding between the side chain of the aspartic acid residue and the guanine ring in nucleotide binding specificity. Besides nucleotide binding specificity, the D119N mutation has little or no effect on the interaction of Ha-Ras with SDC25C, SOS1, GAP, or Raf. Neither does it affect the hydrolysis of nucleotide triphosphate. Like xanthine-nucleotide-specific EF-Tu, xanthine-nucleotide-specific Ras and related proteins will be useful tools for elucidating cellular systems containing multiple regulatory GTPases.
我们研究了在第119位发生天冬氨酸到天冬酰胺取代（D119N突变）的c-Ha-Ras。天冬氨酸119是调节性GTP酶家族成员共有的保守NKXD基序的一部分。有人提出这个天冬酰胺残基参与与鸟嘌呤环的直接结合，并决定鸟嘌呤核苷酸的结合特异性。发现D119N突变会将Ha-Ras的核苷酸特异性从鸟嘌呤改变为黄嘌呤，这一观察结果直接支持了天冬氨酸残基侧链与鸟嘌呤环之间的氢键在核苷酸结合特异性中的重要作用。除了核苷酸结合特异性外，D119N突变对Ha-Ras与SDC25C、SOS1、GAP或Raf的相互作用几乎没有影响。它也不影响三磷酸核苷酸的水解。与黄嘌呤核苷酸特异性的EF-Tu一样，黄嘌呤核苷酸特异性的Ras及相关蛋白将成为阐明含有多种调节性GTP酶的细胞系统的有用工具。
