Influence of pH on the partition of glucose-6-phosphate dehydrogenase and hexokinase in aqueous two-phase system

Silva, Daniel Pereira daPontes, Marcela Zanella RibeiroSouza, Maria Aparecida deVitolo, MicheleSilva, João Batista de Almeida ePessoa-Junior, Adalberto

Glucose-6-phosphate dehydrogenase (G6PDH) and hexokinase (HK) are important enzymes used in biochemical and medical studies and in several analytical methods. Aqueous two-phase system (ATPS) formed by a polymer solution and an electrolyte solution provides a method for the separation and purification of enzymes with several advantages, including biocompatibility and easy scale up of the process. In this work, the effects of different pH values on the storage stability and partitioning behavior (K, partition coefficient) of the enzymes G6PDH and HK from baker's yeast extract were investigated in ATPS. The results, obtained from the 17.5 percent PEG 400 : 15.0 percent phosphate system, showed that when the pH was increased from 5.0 to 8.8, the K HK increased 26-fold and the K G6PDH 2.2-fold. In the 20.0 percent PEG 1500 : 17.5 percent phosphate system, the K HK and K G6PDH increased 13 and 1.2-fold, when the pH value was increased from 3.8 to 8.8, respectively. This leads to the conclusion that the partition coefficient for both enzymes is favored by high pH values. A statistical analysis of the results was conducted to confirm this conclusion