The allosteric activator Mg-ATP modifies the quaternary structure of the R-state of Escherichia coli aspartate transcarbamylase without altering the T ⇆ R equilibrium

被引:21
作者
Fetler, L
Vachette, P
机构
[1] Univ Paris 11, LURE, CNRS, CEA,MER, F-91898 Orsay, France
[2] Univ Paris 06, Lab Biochim Signaux Regulateurs Cellulaires & Mol, CNRS, UMR 7631, F-75006 Paris, France
关键词
allostery; ATCase; small-angle X-ray scattering; conformational change; enzyme regulation;
D O I
10.1006/jmbi.2001.4681
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The allosteric enzyme aspartate transcarbamylase from Escherichia coli (ATCase) displays regulatory properties that involve various conformational changes, including a large quaternary structure rearrangement. This' entails a major change in its solution X-ray scattering curve upon binding substrate analogues. We show here that, in the presence of the nucleotide effector ATP, known to stimulate the enzyme activity, the scattering profiles show a marked dependence on the metal bound to Am. Whereas ATP has no major effect on the scattering pattern of ATCase, a saturating concentration of Mg-ATP notably modifies the scattering profile of the enzyme, either in the absence or in the presence of the bisubstrate analogue N-(phosphonacetyl)-L-aspartate (PALA). The transition with PALA in the presence of this metal-nucleotide complex remains concerted. Furthermore, Mg-ATP, as already observed with Am, has no detectable direct effect on the T to R transition. The experimental scattering curves in the presence of Mg-ATP were fitted by a modeling approach using rigid body movements of the regulatory subunits and the catalytic trimers in the crystal structures. While the differences observed in the T-state in the presence of Mg-ATP are essentially attributed to the binding per se of the nucleotide, the solution structure of the R-state complexed to Mg-Am is even more extended along the 3-fold axis than the previously described R solution structure, which is already more stretched out along the same axis than the crystal R structure. Based on the crystal structure of the enzyme in the R-state complexed with free ATP, a proposal is made to account for the effect of magnesium. (C) 2001 Academic Press.
引用
收藏
页码:817 / 832
页数:16
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