Braxton B L, Mullins L S, Raushel F M, Reinhart G D
Department of Biochemistry and Biophysics, Texas A&M University, College Station 77843-2128, USA.
Biochemistry. 1999 Feb 2;38(5):1394-401. doi: 10.1021/bi982097w.
A linked-function analysis of the allosteric responsiveness of carbamoyl phosphate synthetase (CPS) from E. coli was performed by following the ATP synthesis reaction at low carbamoyl phosphate concentration. All three allosteric ligands, ornithine, UMP, and IMP, act by modifying the affinity of CPS for the substrate MgADP. Individually ornithine strongly promotes, and UMP strongly antagonizes, the binding of MgADP. IMP causes only a slight inhibition at 25 degreesC. When both ornithine and UMP were varied, models which presume a mutually exclusive binding relationship between these ligands do not fit the data as well as does one which allows both ligands (and substrate) to bind simultaneously. The same result was obtained with ornithine and IMP. By contrast, the actions of UMP and IMP together must be explained with a competitive model, consistent with previous reports that UMP and IMP bind to the same site. When ornithine is bound to the enzyme, its activation dominates the effects when either UMP or IMP is also bound. The relationship of this observation to the structure of CPS is discussed.
通过在低氨甲酰磷酸浓度下跟踪ATP合成反应，对来自大肠杆菌的氨甲酰磷酸合成酶（CPS）的变构反应性进行了关联功能分析。所有三种变构配体，即鸟氨酸、UMP和IMP，都是通过改变CPS对底物MgADP的亲和力来发挥作用的。单独来看，鸟氨酸强烈促进，而UMP强烈拮抗MgADP的结合。IMP在25℃时仅引起轻微抑制。当同时改变鸟氨酸和UMP时，假定这些配体之间存在互斥结合关系的模型不如允许两种配体（和底物）同时结合的模型拟合数据。鸟氨酸和IMP同时存在时也得到了相同的结果。相比之下，UMP和IMP共同作用必须用竞争模型来解释，这与之前报道的UMP和IMP结合到同一位点一致。当鸟氨酸与酶结合时，其激活作用在UMP或IMP也结合时占主导地位。讨论了这一观察结果与CPS结构的关系。
