KINETIC-STUDIES OF FR-1, A GROWTH FACTOR-INDUCIBLE ALDO-KETO REDUCTASE

Murine fibroblasts cultured in the presence of fibroblast growth facto r-1 express relatively high levels of FR-1, a similar to 36 kDa protei n related to the aldo-keto reductase superfamily [Donohue, P. J., Albe rts, G. F., Hampton, B. S., Winkles, J.A. (1994) J. Biol. Chem. 269, 8 604-8609]. While the crystal structure of FR-1 shows striking homology with:human aldose reductase [Wilson, D. K., Nakano, T., Petrash, J. M ., Quiocho, F. A. (1995) Biochemistry 34, 14323-14330], an enzyme link ed to the pathogenesis of diabetic complications, the physiological ro le of FR-1 is not known. We show that FR-1 is capable of reducing a br oad range of aromatic and aliphatic aldehydes, :including the abundant and highly reactive lipid-derived aldehyde 4-hydroxy-2-nonenal (HNE; K-m approximate to 9 mu M). However, in the absence of coenzyme, HNE c aused a time-dependent inactivation of FR-1. Results from electrospray ionization-mass spectrometry and Edman-degradation of peptides derive d from HNE-modified FR-1 were consistent with formation of a Michael a dduct at Cys298. This was confirmed with a C298S mutant, which was res istant to HNE-induced inactivation. Since steady-state K-m values dete rmined with alkanals, alpha,beta-unsaturated alkenals, alkadienals, an d 4-hydroxyalkenals fall within their physiological concentrations, li pid-derived aldehydes appear to be potential in vivo substrates for FR -1.