Partition behavior and partial purification of hexokinase in aqueous two-phase polyethylene glycol/citrate systems.
This study dealt with the partition behavior and partial purification of hexokinase (HK) from baker's yeast by liquid-liquid extraction using aqueous two-phase polyethylene glycol (PEG)/citrate systems. First, we investigated the effect of agitation type (vortex and 8 rpm rotation) on the stability of the system, and then the effects of sodium citrate concentration, PEG concentration, and molar mass of PEG on the partition coefficient of this enzyme by using a 25 factorial experimental design. The results of this factorial experiment showed the possibility of a partial purification of HK by using two extraction steps, since the enzyme preferentially migrated to the top phase and the total proteins (mainly contaminants) remained in the bottom phase. The purification factor (PurTOP) of the enzyme in the top phase was 1.87, and the partition coefficient of the total proteins (KProt) was 0.47.
Department of Biochemical and Pharmaceutical Technology, Faculty of Pharmacy, University of São Paulo, Av. Prof. Lineu Prestes, 580, B.16. 05508-900, São Paulo, SP, Brazil.
George G G
GG
Oliveira
Daniel P
DP
Silva
Inês Conceição
IC
Roberto
Michele
M
Vitolo
Adalberto
A
Pessoa
D002951:Citrates; D004795:Enzyme Stability; D006593:Hexokinase / Q000302:isolation & purification* / Q000378:metabolism*; D007202:Indicators and Reagents; D007700:Kinetics; D008970:Molecular Weight; D011092:Polyethylene Glycols; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D000077559:Sodium Citrate; D014867:Water
D002951:Citrates; D007202:Indicators and Reagents; D014867:Water; D000077559:Sodium Citrate; D011092:Polyethylene Glycols; D006593:Hexokinase