Figure 4. Phenotypic profiling predicts the cholesterol catabolic pathway.

Proposed chemical transformations necessary for cholesterol degradation are divided into sterol ring cleavage and opening (A) and side-chain degradation (B). Enzymes known to or predicted to function at each step are noted with their respective fold change (sequence reads in cholesterol/sequence reads in glycerol) below. Dotted lines represent spontaneous reactions. We did not find enzymes with marked * to be significantly required for growth on cholesterol. Enzymes marked with ** indicate those where we are unable to report significance due to insufficient data. The specific β-oxidation steps mediated by fadE28 and fadE29 are difficult to predict based on their homology to testosterone degrading enzymes and therefore, we have assigned them to both equally likely steps.