Oxidized galectin-1 promotes axonal regeneration in peripheral nerves but does not possess lectin properties

Galectin-1 has recently been identified as a factor that regulates initial axonal growth in peripheral nerves after axotomy. Although galectin-1 is a well-known beta-galactoside-binding lectin, its potential to promote axonal regeneration as a lectin has not been reported. It is essential that the p rocess of initial repair in peripheral nerves after axotomy is well clarifi ed. We therefore undertook to investigate the relation between the structur e and axonal regeneration-promoting activity of galectin-1. Recombinant hum an galectin-1 secreted into the culture supernatant of transfected COS1 cel ls (rhGAL-1/COS1) was purified under nonreducing conditions and subjected t o structural analysis. Mass spectrometric analysis of peptide fragments fro m rhGAL-1/COS1 revealed that the secreted protein exists as an oxidized for m containing three intramolecular disulfide bonds (Cys2-Cys130, Cys16-Cys88 and Cys42-Cys60). Recombinant human galectin-1 (rhGAL-1) and a galectin-1 mutant in which all six cysteine residues were replaced by serine (CSGAL-1) were expressed in and purified from Escherichia coli for further analysis; the purified rhGAL-1 was subjected to oxidation, which induced the same pa ttern of disulfide linkages as that observed in rhGAL-1/COS1. Oxidized rhGA L-1 enhanced axonal regeneration from the transected nerve sites of adult r at dorsal root ganglion explants with associated nerve stumps (5.0-5000 pg. mL(-1)), but it lacked lectin activity. In contrast, CSGAL-1 induced hemagg lutination of rabbit erythrocytes but lacked axonal regeneration-promoting activity. These results indicate that galectin-1 promotes axonal regenerati on only in the oxidized form containing three intramolecular disulfide bond s, not in the reduced form which exhibits lectin activity.