荧光素酶在PEG/盐系统中分配系数的研究

The Partition Coefficient of Luciferase from Photobacteria in the PEG/salt Two Aqueous Phase System

海洋发光细菌产生的荧光素酶在PEG/盐双水相系统中的分配系数(K)的对数值与PEG分子量成线性关系。低分子量PEG亲水性较强,有利于酶分配于PEG上相中;而高分子量PEG疏水性较强,有利于酶分配在下相盐溶液中,PEG/三价盐与PEG/二价盐系统不同,前者有一折点。在使用单一分子量PEG组成的PEG/硫酸铵系统中,K值随系统中使用的硫酸铵浓度增加而增加,硫酸铵达到一定浓度后,其浓度继续增加对K值的增加影响减小;但在两种不同分子量的PEG混合物组成的PEG/硫酸铵系统中,K值随硫酸铵浓度的增加有一最小值,在此硫酸铵浓度之外,增加或减小硫酸铵浓度,K值均会增加。在PEG/硫酸铵系统中,荧光素酶的K值随系统的pH增加略有提高,但变化不大。而在PEG/磷酸盐系统中,在pH6.4时,K值最小,pH高或低于此值,K值均增加。系统的温度变化对K值影响很小。

The relationship between the logarithmic partition coefficient(K) of luciferase of photobacteria and PEG MW in the PEG/salt two aqueous phase system shown a linear function. Hydrophilic PEG with low MW facilitated the partition of luciferase into the top PEG-riching phase and give a higher K value, while the high MW one, due to its hydrophobic characterstic, made the enzyme partition more into the bottom salt-riching phase, and a lower K value. In the PEG/trivalent salt system, such as phosphate and citrate, there is a turn-point on the linear relation between the log K and the PEG MW, but it is never appeared if a divalent salt such as sulfate, succinate or tartrate is used in the system.If the system was compased of PEG with homogenous MW and ammonium sulfate, the K value would be increased with the increment of the salt concentration, but after the salt concentration had reached at certain level, the K value would not be influenced. If two kind of PEGs with different MW were used in this system a minimal K value would be appeared at a certain ammonium sulfate concentration, and the K value would be raised when the salt concentration was either increased or decreased.Neither the proportion of the two kind of PEGs nor their total con- centration used in the system would pose any impaction the above patterns, although the K value would have a corresponding change.In PEG/ammonium sulfate system, the K value was slightly increased with the increase of the system pH, while in the PEG/phosphate system, a minimal K value appeares at pH6.4 and it would go up when the system pH was either increased or decreased. The effect of system temperature on K value was very sligbt.