Characterization of patatin esterase activity in AOT-isooctane reverse micelles.
Patatin is a family of glycoproteins that accounts for 30-40% of the total soluble protein in potato (Solanum tuberosum L.) tubers. This protein has been reported not only to serve as a storage protein but also to exhibit lipid acyl hydrolase (LAH) activity. In this study patatin is characterized in AOT-isooctane reverse micelles. The influence on the enzymatic activity of characteristic parameters of reverse micelles, w(o) (= H(2)O/AOT), and the percentage of H(2)O, theta, were investigated. The results obtained show that patatin esterase activity varies with w(o) but remains constant throughout the range of theta values studied. The variation with w(o) showed that the activity follows an S-shaped behavior pattern, reaching a maximum at about w(o) = 20 for 2% H(2)O. Patatin esterase activity was compared with p-nitrophenyl (PNP) fatty acid esters of different chain lengths. The activity was much higher for PNP-caprylate. The pH optimum was 6.0, different from the value obtained when patatin esterase activity was measured in mixed micelle systems. The optimal temperature was 35 degrees C, above which the activity decreased to almost zero. The kinetic parameters were also evaluated (K(m) = 10 mM, V(m) = 158 microM/min, V(m)/K(m) = 15.8 x 10(-3) min(-1)). This paper shows the suitability of reverse micelles for measuring patatin esterase activity, since it allows the study of the enzyme in similar conditions to that prevailing in vivo.
Jiménez
M
M
Departamento de Bioquímica y Biología Molecular A, Universidad de Murcia, Espinardo 30071, Murcia, Spain. mja@um.es
Escribano
J
J
Gandía-Herrero
F
F
Chazarra
S
S
Cabanes
J
J
García-Carmona
F
F
Pérez-Gilabert
M
M
D002265:Carboxylic Ester Hydrolases / Q000737:chemistry*; D004143:Dioctyl Sulfosuccinic Acid / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008823:Micelles; D009795:Octanes / Q000737:chemistry*; D010940:Plant Proteins / Q000737:chemistry*; D013379:Substrate Specificity; D013696:Temperature
D008823:Micelles; D009795:Octanes; D010940:Plant Proteins; C043222:patatin protein, Solanum tuberosum; D004143:Dioctyl Sulfosuccinic Acid; D002265:Carboxylic Ester Hydrolases; C045798:2,2,4-trimethylpentane