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Upon sonication, the antimicrobial and antineoplastic compound dequalinium forms vesicles (DQAsomes, Weissig et al., 1998). Dequalinium (1,1'-(1,10-decamethylene-bis-[aminoquinaldinium])-chloride) was shown to be a fluorophore with an emission maximum at 366 nm. Addition of DNA results in a characteristic quenching of its intrinsic fluorescence. After density gradient centrifugation a band of dequalinium (DQA) tightly associated with DNA is located between the DNA and DQA bands. DQA/DNA-complexes containing plasmid DNA at a molar ratio of DQA/DNA 6:1 are completely protected against DNase activity. Addition of negatively-charged lipids release intact DNA in the same manner as from cationic lipid/DNA complexes. As regards biological effects, DQAsomes show a differential cytotoxicity for normal and sarcoma cell lines. In vitro incubation with fluorescein-labeled oligodeoxynucleotides (5'-fluorescein-[GATC]5) showed an increased uptake of the tagged oligodeoxynucleotide if complexed with dequalinium. We hypothesize that the DQA/DNA complexes are well-suited for 'DQAsomal gene transfer' in vitro and in vivo. Noteworthy, they display an intrinsic antitumor activity manifested by differential cytotoxicity for normal and sarcoma cells.

The solvolytic detachment of leucine aminopeptidase from Sepharose-enzyme conjugates with multiple and single anchoring bonds has been studied under a variety of conditions by radiochemical and enzymological methods. The release of the single-point-fixed conjugate could be described by a leakage function, derived previously, yielding the first-order rate constant of the cleavage of the enzyme-matrix bond. The nucleophile hydroxylamine increased the detachment rate considerably. The release of the immobilized enzyme was incomplcte in all experiments even after prolonged times. The enzyme leakage from multipoint-attached conjugates was still high enough to prohibit a long-term use of such preparations in routine work at room temperature.Although the first covalent attachments of biologically active proteins to insoluble carriers date back to the early fifties [1,2], the widespread use of artificially immobilized enzymes had to await the advent of a simple and straightforward method resulting in a high efficiency of binding and the preservation of most of the activity. The introduction of the cyanogen bromide activation of insoluble polysaccharides for the coupling with ligand molecules [3] filled this gap.Recent studies have, however, drawn attention to the fact that this most widely used method suffers from a serious drawback : low-molecular-weight ligands are slowly released from the polymeric support in aqueous surroundings above pH 5 [4-61. This detachment may beashighas 1 %ofthetotalamountofimmobilized ligand per hour, as has been shown with Sepharosebound ~-phenylalanine-4-nitroanilide at pH 7.8 and 25 "C [7]. The chemical mechanism of this relatively facile solvolytic detachment has been elucidated in considerable detail [5], though the surmised anchimeric assistance of neighbouring groups of the carrier gel needs further studies.In the presence of strong nucleophiles the detachment rate is increased considerably so that even multiAbbreviations. Thiol-Sepharose, glutathionine-derivative of Sepharose; Sepharose-aminopeptidase, leucine amino-peptidase covalently bound to Sepharose by a multitude of bonds (CNBr method); Sepharose-S-S-aminopeptidase. leucine amino-peptidase attached to thiol-Sepharose by a disulfide bond. On the other hand it has been demonstrated that derivatives of high stability can be prepared by coupling the multivalent mediators, polylysine, polyornithine or polyvinylamine, to Sepharose [ll].It seems that the possibility of a release of BrCNimmobilized enzymes around the neutral point in the absence of strong nucleophiles has so far been considered negligible and not studied systematically.Some attempts have been made to describe mathematically the leakage of low-molecular-weight ligands from affinity gels [12,13]. However, with immobilized enzymes, where quite a number of heterogeneous bonds between carrier and macromolecular ligand can be anticipated, the assumptions on which the derivations of leakage functions are based will hold only in exceptional cases.So far, the multiplicity and ...

The physiological function of the GPI-anchored ectoenzyme aminopeptidase P (APP) is still elusive. Most researchers suppose that this enzyme inactivates biologically active peptides like bradykinin, neuropeptide tyrosine (NPY) and others (Vanhoof et al., 1995). We demonstrate by immunohistology with a specific antibody raised in rabbits and measurement of enzymatic activity in suspensions and of confluent monolayers on microscopic coverslips ('monolayer kinetics') that APP is a cell surface enzyme (ectoenzyme) of endothelial and lymphoid cells.

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