FIGURE 5.

A, electron microscopy of the phosphatidylcholine liposomes used before (left) and after (middle) hexane addition. The bar represents 200 nm. Lycopene cyclization took place in highly turbid assays in which lycopene (red) was converted into β-carotene (yellow). A standard assay is shown in the absence (Con) and presence of mCrtY after 1 h of incubation. B, shown is the time course of lycopene cyclization using 5 μg of holo-mCrtY isolated according to protein purification method 2 in anaerobic standard assays in the presence of different reductants. When photoreduction was used, the addition of FAD was needed to achieve activity in a concentration-dependent manner, raising the specific activity from 0.14 pmol μg of mCrtY⁻¹ min⁻¹ (20 μm FAD) to 2.2 pmol μg of mCrtY⁻¹ min⁻¹ (400 μm FAD). No addition of FAD was needed when Ti(III) citrate was used as the reductant to achieve an activity of 1.7 pmol μg of mCrtY⁻¹ min⁻¹, but FAD addition (20 μm) was stimulatory, leading to an activity of 2.5 pmol μg of mCrtY⁻¹ min⁻¹. The stimulation achieved by FAD addition indicates the presence of some apo-mCrtY. Filled symbols are for assays carried out in the absence of a reductant both in the presence or absence of FAD, resulting in complete absence of enzymatic activity.