G13A Substitution Affects the Biochemical and Physical Properties of the Elongation Factor 1. A Reduced Intrinsic GTPase Activity Is
详细信息 查看全文
- 作者：Mariorosario Masullo ; Piergiuseppe Cantiello ; Barbara de Paola ; Francesca Catanzano ; Paolo Arcari ; and Vincenzo Bocchini
- 刊名：Biochemistry
- 出版年：2002
- 出版时间：January 15, 2002
- 年：2002
- 卷：41
- 期：2
- 页码：628 - 633
- 全文大小：85K
- 年卷期：v.41,no.2(January 15, 2002)
- ISSN：1520-4995
文摘
The G13A substitution in the G13XXXXGK[T,S] consensus sequence of the elongation factor1
from the archaeon Sulfolobus solfataricus (SsEF-1) was introduced in order to study the reasons forselective differences found in the homologous consensus element AXXXXGK[T,S] of the other elongationfactor EF-2 or EF-G. In a previous work, it was shown that the main effect of the A26G mutation wasthe activation of the intrinsic GTPase of SsEF-2 [De Vendittis, E., Adinolfi, B. S., Amatruda, M. R.,Raimo, G., Masullo, M., and Bocchini, V. (1994) Eur. J. Biochem. 262, 600-605]. In this work, wefound that, compared to the wild-type factor (SsEF-1 wt), G13ASsEF-1 shows (i) a reduced rate of[3H]Phe polymerization that was probably due to its reduced ability to form a ternary complex withheterologous aa-tRNA and (ii) a reduced intrinsic GTPase activity that was stimulated by high concentrationsof NaCl (GTPaseNa) [Masullo, M., De Vendittis, E., and Bocchini, V. (1994) J. Biol. Chem. 269, 20376-20379]. In addition, G13ASsEF-1 showed an increased affinity for GDP and GTP. Surprisingly, thedecreased intrinsic GTPaseNa of G13ASsEF-1 can be partially restored by kirromycin, an effect notfound for SsEF-1 wt. The temperature inducing a 50% denaturation of G13ASsEF-1 was somewhatlower (-5 
C) than that of SsEF-1 wt, and the decrease in its thermophilicity was slightly more accentuated(-10 C). These results indicate that the nature of the residue in position 13 is important for the functionaland physical properties of SsEF-1.
from the archaeon Sulfolobus solfataricus (SsEF-1) was introduced in order to study the reasons forselective differences found in the homologous consensus element AXXXXGK[T,S] of the other elongationfactor EF-2 or EF-G. In a previous work, it was shown that the main effect of the A26G mutation wasthe activation of the intrinsic GTPase of SsEF-2 [De Vendittis, E., Adinolfi, B. S., Amatruda, M. R.,Raimo, G., Masullo, M., and Bocchini, V. (1994) Eur. J. Biochem. 262, 600-605]. In this work, wefound that, compared to the wild-type factor (SsEF-1 wt), G13ASsEF-1 shows (i) a reduced rate of[3H]Phe polymerization that was probably due to its reduced ability to form a ternary complex withheterologous aa-tRNA and (ii) a reduced intrinsic GTPase activity that was stimulated by high concentrationsof NaCl (GTPaseNa) [Masullo, M., De Vendittis, E., and Bocchini, V. (1994) J. Biol. Chem. 269, 20376-20379]. In addition, G13ASsEF-1 showed an increased affinity for GDP and GTP. Surprisingly, thedecreased intrinsic GTPaseNa of G13ASsEF-1 can be partially restored by kirromycin, an effect notfound for SsEF-1 wt. The temperature inducing a 50% denaturation of G13ASsEF-1 was somewhatlower (-5 C) than that of SsEF-1 wt, and the decrease in its thermophilicity was slightly more accentuated(-10 C). These results indicate that the nature of the residue in position 13 is important for the functionaland physical properties of SsEF-1.