Biochemical and biological characterization of a new oxidized avidin with enhanced tissue binding properties.
Chicken avidin and bacterial streptavidin are widely employed in vitro for their capacity to bind biotin, but their pharmacokinetics and immunological properties are not always optimal, thereby limiting their use in medical treatments. Here we investigate the biochemical and biological properties of a new modified avidin, obtained by ligand-assisted sodium periodate oxidation of avidin. This method allows protection of biotin-binding sites of avidin from inactivation caused by the oxidation step and delay of avidin clearance from injected tissue by generation of aldehyde groups from avidin carbohydrate moieties. Oxidized avidin shows spectroscopic properties similar to that of native avidin, indicating that tryptophan residues are spared from oxidation damage. In strict agreement with these results, circular dichroism and isothermal titration calorimetry analyses confirm that the ligand-assisted oxidation preserves the avidin protein structure and its biotin binding capacity. In vitro cell binding and in vivo tissue residence experiments demonstrate that aldehyde groups provide oxidized avidin the property to bind cellular and interstitial protein amino groups through Schiff's base formation, resulting in a tissue half-life of 2 weeks, compared with 2 h of native avidin. In addition, the efficient uptake of the intravenously injected (111)In-BiotinDOTA (ST2210) in the site previously treated with modified avidin underlines that tissue-bound oxidized avidin retains its biotin binding capacity in vivo. The results presented here indicate that oxidized avidin could be employed to create a stable artificial receptor in diseased tissues for the targeting of biotinylated therapeutics.
Verdoliva
Antonio
A
Research and Development Department, Tecnogen SpA, Piana di Monte Verna, 81013 Caserta, Italy.
Bellofiore
Piero
P
Rivieccio
Vincenzo
V
Catello
Sergio
S
Colombo
Maurizio
M
Albertoni
Claudio
C
Rosi
Antonio
A
Leoni
Barbara
B
Anastasi
Anna Maria
AM
De Santis
Rita
R
D000447:Aldehydes / Q000737:chemistry; D000818:Animals; D001360:Avidin / Q000737:chemistry*; D001671:Biochemistry / Q000379:methods*; D019921:Biotinylation / Q000379:methods; D002645:Chickens; D005260:Female; D005434:Flow Cytometry; D008024:Ligands; D051379:Mice; D008807:Mice, Inbred BALB C; D010100:Oxygen / Q000737:chemistry; D010504:Periodic Acid / Q000737:chemistry; D011485:Protein Binding; D013056:Spectrophotometry, Ultraviolet / Q000379:methods
D000447:Aldehydes; D008024:Ligands; D010504:Periodic Acid; D001360:Avidin; C009288:metaperiodate; D010100:Oxygen