Cortese J D, Vidal J C
Arch Biochem Biophys. 1983 Jul 1;224(1):351-7. doi: 10.1016/0003-9861(83)90219-9.
D-beta-hydroxybutyrate dehydrogenase, purified as soluble, lipid-free apoenzyme (inactive) from rat liver mitochondria can be reactivated by the short-chain dihexanoyl, diheptanoyl, and dioctanoyl lecithins at the monomeric state, upon formation of a reversible enzyme-lecithin complex. Previous studies with these lecithins suggested that reactivation of the apoenzyme requires the simultaneous occupation of two identical, noninteracting lecithin binding sites via a rapid equilibrium random mechanism. The short-chain lecithins exhibited similar reactivating capacities, differing only in their affinities towards the enzyme. In order to further test that model, the reactivation of the apoenzyme was studied when two or three short-chain lecithins were simultaneously present in the reaction medium. The initial velocities were measured either as a function of the concentration of one lecithin while the other(s) were kept constant, or as a function of the total phospholipid concentration with mixtures of different lecithins at a constant molar ratio. The pertinent equations were derived on the principles of multiple equilibria with identical, noninteracting sites able to be occupied by any of the different lecithins present in the reaction medium, with the doubly occupied enzyme as the only active species. In agreement with the above-proposed model, the results obtained indicates that the molar fraction of the doubly occupied (active) enzyme species can be calculated from equilibrium considerations and that the maximal attainable with the different short-chain lecithins are similar.
从大鼠肝脏线粒体中纯化得到的D-β-羟基丁酸脱氢酶，以可溶性、无脂质的脱辅基酶（无活性）形式存在，在形成可逆的酶-卵磷脂复合物时，可被单体状态的短链二己酰基、二庚酰基和二辛酰基卵磷脂重新激活。此前对这些卵磷脂的研究表明，脱辅基酶的重新激活需要通过快速平衡随机机制同时占据两个相同的、不相互作用的卵磷脂结合位点。短链卵磷脂表现出相似的重新激活能力，只是它们对酶的亲和力有所不同。为了进一步验证该模型，研究了在反应介质中同时存在两种或三种短链卵磷脂时脱辅基酶的重新激活情况。初始速度的测量方式有两种：一种是在其他卵磷脂浓度保持恒定的情况下，作为一种卵磷脂浓度的函数进行测量；另一种是在不同卵磷脂以恒定摩尔比混合时，作为总磷脂浓度的函数进行测量。根据多重平衡原理推导了相关方程，其中相同的、不相互作用的位点能够被反应介质中存在的任何一种不同卵磷脂占据，而只有双重占据的酶是唯一的活性物种。与上述提出的模型一致，所得结果表明，双重占据（活性）酶物种的摩尔分数可以通过平衡考虑来计算，并且不同短链卵磷脂所能达到的最大值相似。
