Abstract
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.
Highlights
Addition, the use of avidin to neutralize the anticoagulant activity of idrabiotaprinux by increasing the plasma clearance of the idrabiotaprinux-avidin complex has been demonstrated [7]
The results indicate that the hydroxyazobenzene-2Ј-carboxylic acid (HABA)-assisted oxidation substantially preserves the structure and the biotin binding property of native avidin, which acquires the capacity to reside within injected tissues for weeks as a consequence of the chemical reaction of aldehyde groups with tissue protein amino groups by Schiff’s base formation
Cell Binding and Tissue Kinetics of Avidin and Streptavidin— Looking for a stable receptor for radiolabeled biotin in tissues to be exposed to radiotherapy, preliminary experiments were performed to investigate the in vitro cell binding and in vivo tissue residence and efficiency in capturing the ST2210 radioactive biotin of both reference molecules avidin and streptavidin
Summary
Addition, the use of avidin to neutralize the anticoagulant activity of idrabiotaprinux by increasing the plasma clearance of the idrabiotaprinux-avidin complex has been demonstrated [7]. Enhanced Tissue Binding Properties of Oxidized Avidin low affinity ligand 4-hydroxyazobenzene-2Ј-carboxylic acid (HABA)2 [36] This reaction produces reactive aldehyde groups (CHO) that are virtually inert at acidic pH and, when injected in a tissue, because of the physiological pH, react with NH2 groups on tissue proteins, delaying clearance of such modified avidin, named OXavidinHABA. The results indicate that the HABA-assisted oxidation substantially preserves the structure and the biotin binding property of native avidin, which acquires the capacity to reside within injected tissues for weeks as a consequence of the chemical reaction of aldehyde groups with tissue protein amino groups by Schiff’s base formation. It could be employed for other medical applications where a stable artificial receptor in diseased tissues could be envisaged to direct biotinylated therapeutics
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