Hauryliuk Vasili, Zavialov Andrey, Kisselev Lev, Ehrenberg Måns
Department of Cell and Molecular Biology, Molecular Biology Program, BMC, Box 596, Uppsala University, 75124, Sweden.
Biochimie. 2006 Jul;88(7):747-57. doi: 10.1016/j.biochi.2006.06.001. Epub 2006 Jun 15.
In eukaryotes, termination of mRNA translation is triggered by the essential polypeptide chain release factors eRF1, recognizing all three stop codons, and eRF3, a member of the GTPase superfamily with a role that has remained opaque. We have studied the kinetic and thermodynamic parameters of the interactions between eRF3 and GTP, GDP and the non-hydrolysable GTP analogue GDPNP in the presence (K(D)(GDP)=1.3+/-0.2 muM, K(D)(GTP) approximately 200 muM and K(D)(GDPNP)>160 muM) as well as absence (K(D)(GDP)=1.9+/-0.3 muM, K(D)(GTP) 0.7+/-0.2 muM and K(D)(GDPNP) approximately 200 muM) of eRF1. From the present data we propose that (i) free eRF3 has a strong preference to bind GDP compared to GTP (ii) eRF3 in complex with eRF1 has much stronger affinity to GTP than free eRF3 (iii) eRF3 in complex with PABP has weak affinity to GTP (iv) eRF3 in complex with eRF1 does not have strong affinity to GDPNP, implying that GDPNP is a poor analogue of GTP for eRF3 binding.
在真核生物中，mRNA翻译的终止由必需的多肽链释放因子eRF1触发，eRF1可识别所有三种终止密码子，以及eRF3，它是GTPase超家族的成员，其作用一直不明确。我们研究了在有（K(D)(GDP)=1.3±0.2 μM，K(D)(GTP)约200 μM，K(D)(GDPNP)>160 μM）和无（K(D)(GDP)=1.9±0.3 μM，K(D)(GTP) 0.7±0.2 μM，K(D)(GDPNP)约200 μM）eRF1的情况下，eRF3与GTP、GDP和不可水解的GTP类似物GDPNP之间相互作用的动力学和热力学参数。根据目前的数据，我们提出：(i) 与GTP相比，游离的eRF3对结合GDP有强烈偏好；(ii) 与eRF1形成复合物的eRF3对GTP的亲和力比游离的eRF3强得多；(iii) 与PABP形成复合物的eRF3对GTP的亲和力较弱；(iv) 与eRF1形成复合物的eRF3对GDPNP没有很强的亲和力，这意味着GDPNP是eRF3结合的GTP的不良类似物。
