Table 2.5.
RA biosynthesis catalyzed by microsomes (RDH activity) and a cytosolic fraction (RALDH activity) using CRBP1-retinol as substrate1
| RA generated (pmol ± SD)
|
||||||||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|
| Addition | 0 | 0 | 0 | 0 | 0 | 0 | 0 | ~2 | 5±4 | 9±5 | 20±2 | 49±4 |
| μsomes | + | + | + | − | − | − | + | − | + | + | + | + |
| P1 | − | − | − | + | + | + | + | + | − | + | + | + |
| NAD+ | − | + | − | − | + | − | − | + | + | + | − | + |
| NADP+ | − | − | + | − | − | + | − | + | + | − | + | + |
1
A total of 7.5 μM CRBP1 was used with 5 μM retinol to insure minimal free retinol [212]. With a kd of 3 nM, the unbound retinol concentration was 6 nM, whereas the holo-CRBP1 concentration was 4994 nM. The liver cytosol fraction P1 with RALDH activity was isolated by anion-exchange chromatography