Oxidative radioiodination damage to human lactoferrin

Abstract

Oxidative iodination of human lactoferrin (Lf) as commonly performed by using the chloramine-T, the Iodogen or the lactoperoxidase method produces an unreliable tracer protein because of excessive and heterogeneous polymer formation. Before iodination a minor tetramer fraction may be demonstrable in iron-saturated Lf only. Iodination-induced polymerization of iron-poor as well as iron-saturated Lf occurs independently of the presence or absence of 10 mM-EDTA and the 125I-/Lf molar ratio used for iodination. 125I-Lf polymers are mainly covalently linked, as suggested by the lack of substantial dissociation in SDS/polyacrylamide-gel electrophoresis. Damage to the 125I-Lf monomer may be another consequence of oxidative iodination. This is demonstrated in SDS/polyacrylamide-gel electrophoresis where 50% of the radioactivity of apparently normal monomer (Mr 75,000) is displaced to a lower-Mr region (30,000-67,000) after reduction with dithiothreitol. Non-oxidative iodination by the Bolton-Hunter technique produces an antigenetically stable tracer that is not being subjected to polymerization and monomer degradation as judged by high-performance gel chromatography and SDS/polyacrylamide-gel electrophoresis with and without dithiothreitol treatment. It is concluded that oxidation in itself leads to covalent non-disulphide cross-linking between human Lf molecules and, possibly, to intramolecular peptide-bond breaking becoming unmasked under reducing conditions. In biological experiments with human 125I-Lf this problem should be carefully considered.

Multivalent Metal-Induced Iron Acquisition from Transferrin and Lactoferrin by Myeloid Cells

“…Cell-associated 125 I was then assessed by counting the cell pellet in a gamma counter. It has previously been reported that the binding properties of lactoferrin can be altered by iodination procedures (54). However, the iodination procedure used in our work had a negligible effect on binding of either protein to the HL-60 cell surface, as demonstrated by the following results.…”

Transport of lactoferrin from blood to bile in the rat

“…However, unlike ordinary endosomes, the vesicles transporting lactoferrin to the bile seem ill-equipped to strip iron off this protein. (The small amount of iron lost during transport to the bile could have resulted from oxidative damage to, and ensuing weaker iron binding by, a portion of the iodinated protein [35].) The impression thus gained is that they may represent a distinct type of transport facility.…”