A study was made of the regulation of three enzymes that act sequentially in the metabolism of thymidine in Escherichia coli K-12. Under a variety of conditions, two of the enzymes, thymidine phosphorylase and deoxyribose-5-phosphate aldolase, were found to be synthesized coordinately. However, the third enzyme, phosphodeoxyribomutase, was synthesized noncoordinately with the other two enzymes under the same conditions. In addition, the mutase could be fully induced, whereas basal levels of the phosphorylase and the aldolase were maintained. These findings indicate that two operons comprise the genes concerned with the reversible pathway leading from thymidine to acetaldehyde and glyceraldehyde-3-phosphate. In addition to thymidine, it was found that acetaldehyde was an external inducer of these enzymes. The results of induction experiments performed on wild-type cells and mutants defective in the mutase or the aldolase, with thymidine or acetaldehyde as exogenous inducers, strongly suggest that deoxyribose-5-phosphate is more proximal to the intracellular inducer than is thymidine, deoxyribose-1-phosphate, or acetaldehyde.