Liposomes (lipid vesicles) with sizes of about 100–200
nm
carrying surface-bound (immobilized) water-soluble enzymes are functionalized
molecular compartment systems for possible applications, for example,
as therapeutic materials or as catalytic reaction units for running
reactions in aqueous mediain vitro. One way of covalently
attaching enzyme molecules under mild conditions in a controlled way
to the surface of preformed liposomes is to apply the spectrophotometrically
traceable bis-aryl hydrazone (BAH) bond between the liposome and the
enzyme molecules of interest. Using bovine carbonic anhydrase (BCA),
an aqueous dispersion of liposome-BAH-BCA − conjugates of defined
composition was prepared. The liposomes used consisted of 1,2-dioleoyl-sn-glycero-3-phosphocholine (DOPC),N-(methylpolyoxyethylene
oxycarbonyl)-1,2-distearoyl-sn-glycero-3-phosphoethanolamine
(DSPE-PEG), andN-(aminopropylpolyoxyethylene oxycarbonyl)-1,2-distearoyl-sn-glycero-3-phosphoethanolamine (DSPE-PEG-NH2). The amino group of some of the DSPE-PEG-NH2molecules
present in the liposomes were converted into an aromatic aldehyde,
which (after purification) reacted with (purified) BCA molecules that
had on their surface on average one acetone protected aromatic hydrazine.
After purification of the liposome-BAH-BCA conjugate dispersion obtained,
it was characterized in terms of (i) BCA activity, (ii) overall BCA
structure, and (iii) storage stability. For an average liposome of
138 nm diameter, about 1200 BCA molecules were attached to the outer
liposome surface. Liposomally bound BCA was found to exhibit (i) similar
catalytic activity at 25 °C and (ii) similar storage stability
when stored in a dispersed state in aqueous solution at 4 °C
as free BCA. Measurements at 5 °C clearly showed that liposome-BAH-BCA
is able to catalyze the hydration of carbon dioxide to hydrogen carbonate.