APOBEC-1 INTERACTS WITH A 65-KDA COMPLEMENTING PROTEIN TO EDIT APOLIPOPROTEIN-B MESSENGER-RNA IN-VITRO

The editing of apolipoprotein-B (apoB) mRNA involves the deamination o f cytidine at nucleotide 6666 to uridine, The catalytic subunit of the editing enzyme, apobec-1, is a cytidine deaminase that requires other unidentified proteins to edit apoB mRNA in vitro. We partially purifi ed an activity from baboon kidney that functionally complements apobec -1, The complementing activity was protease-sensitive and micrococcal nuclease-resistant, had a native molecular mass of 65 +/- 10 kDa on si ze exclusion chromatography, and sedimented at 4.5 S in glycerol gradi ents, Purified recombinant His,tagged apobec-1 immobilized on beads de pleted >90% of the complementing activity hom partially purified extra cts, These beads edited apoB mRNA in vitro in the absence of exogenous apobec-1 or complementing activity, A functional holoenzyme containin g apobec-1 and the complementing activity was eluted from the apobec-1 -affinity resin using 0.5 nn imidazole, whereas buffer containing 0.4 M KCl eluted only the complementing activity, The carboxyl-terminal 59 amino acids of apobec-1 were not required for interaction with the co mplementing activity in vitro. Our results demonstrate that the comple menting protein interacts directly with apobec-1 in the absence of apo B mRNA.