Aarhus University Seal / Aarhus Universitets segl

Characterization of lipase in reversed micelles formulated by Cibacron Blue F-3GA modified Span 85

Publikation: Bidrag til tidsskrift/Konferencebidrag i tidsskrift /Bidrag til avisTidsskriftartikelForskningpeer review

  • Dong Hao Zhang, Tianjin University, Kina
  • Zheng Guo
  • Yan Sun, Tianjin University, Kina
  • Molekylærbiologisk Institut
Sorbitan trioleate (Span 85) modified by Cibacron Blue F-3GA (CB) was prepared and used as
an affinity surfactant to formulate a reversed micellar system for Candida rugosa lipase (CRL)
solubilization. The system was characterized and evaluated by employing CRL-catalyzed
hydrolysis of olive oil as a model reaction. The micellar hydrodynamic radius results reflected,
to some extent, the redistribution of surfactant and water after enzyme addition, and the correlation
between surfactant formulation, water content (W0), micellar size, and enzyme activity. An
adequate modification density of CB was found to be important for the reversed micelles to
retain enough hydration capacity and achieve high enzyme activity. Compared with the results
in AOT-based reversed micelles, CRL in this micellar system exhibited a different activity behavior
versus W0. The optimal pH and temperature of the encapsulated lipase remained unchanged,
but the apparent activity was significantly higher than that of the native enzyme in bulk solution.
Kinetic studies indicated that the encapsulated lipase in the reversed micelles of CB-formulated
Span 85 followed the Michaelis-Menten equation. The Michaelis constant was found to decrease
with increasing surfactant concentration, suggesting an increase of the enzyme affinity for the
substrate. Stability of the lipase in the reversed micelles was negatively correlated to W0.
Introduction
Reversed micelles are nanometer-scale transparent aggregates
of water and surfactant dispersed in a bulk apolar solvent where
the surfactant molecules are arranged with their polar head
toward the water pool and hydrophobic tail in contact with bulk
apolar solvent (1). This has been associated to the idea that
reversed micelles mimic those naturally occurring proteins
located in the membrane with both aqueous and nonaqueous
domains (2). The enzyme confined to the water pool of the
reversed micelles is therefore supposed to be capable of retaining
its conformation and activity, which is of particular importance
for those enzymes with higher optimum working water activity
(aw) (3). The microencapsulated enzymes are also assumed to
be sheltered and protected from the detrimental effects of solvent
(2, 4). Compared with other interfacial media, reversed micelles
give a larger interfacial area, facilitating efficient contact
between enzyme and substrate (5). Previous work has also
shown that reversed micellar technology could increase the
apparent activity of enzymes and even yield superactivity for
specific enzymes (6). Lipases are ubiquitous enzymes that could
find many important applications (7, 8). The natural substrate
of lipases is triglyceride, and the majority of lipases are
interfacial activated enzymes (9). This makes lipase an interesting
model enzyme for reversed micelle system studies, in which
lipase-catalyzed
OriginalsprogEngelsk
TidsskriftBiotechnology Progress
Vol/bind23
Nummer1
Sider (fra-til)108-115
ISSN8756-7938
DOI
StatusUdgivet - 2007

Se relationer på Aarhus Universitet Citationsformater

ID: 19548842