Expression and Purification of CRABPs from E. coli

Abstract

Nuclear-retinoid receptors provide a mechanism of retinoid action, but most likely where expressed, the cytosolic cellular retinoic acid-binding proteins (CRABPs) also affect the ability of retinoids to initiate biological signals (1, 2). holo-CRABP I sequesters retinoic acid (RA) with a K _d value that may be <1 nM, and serves as a high-affinity (K _m ~2 nM), efficient substrate of RA metabolism, thereby controlling not only the steady-state concentration of RA, but also its availability (3). The impact of CRABP on retinoid metabolism may extend beyond RA. CRABP binds RA metabolites, including 4-OH-RA, 4-oxo-RA, and several metabolites whose structures have not been identified, and could thereby influence then disposition in vivo (4). For example the elimination t_1/2 of RA in the presence and absence of CRABP I was 35 and 40 min, respectively, in incubations with rat testis microsomes Unbound 4-OH-RA and 4-oxo-RA had elimination t_1/2 values of 40 and 6 min, respectively. In contrast, CRABP-bound 4-OH-RA and CRABP-bound 4-oxo-RA were essentially metabolically inert. This shows that CRABP does not affect all retinoids similarly and that it could have a profound influence on the steadystate concentrations of several retinoids in vivo.