Expression Analysis of <TEX>${eta}$</TEX>-Ketothiolase and Acetoacetyl-CoA Reductase of Rhodobacter sphaeroides

By a sequential action of <TEX>${eta}$</TEX>-ketothiolase and acetoacetyl-CoA reductase, two molecules of acetyl-CoA re converted into D-3-hydroxybutyryl-CoA, a substrate for PHB synthase to form poly-3-hydroxybutyryl-CoA, a substrate for PHB synthase to form poly-3-hydroxybutyrate (PHB) of rhodobacter sphaeroides. The <TEX>${eta}$</TEX>-ketothiolase gene, phbA, and acetoacetyl-CoA reductase gene, phbB, were cloned and analyzed for their expression. Enzyme activities of <TEX>${eta}$</TEX>-ketothiolase and acetoacetyl-CoA reductase showed constitutive levels during aerobic and photoheterotrophic growth of R. sphaeroides. In addition, no difference of each enzyme activity was observed between cells grown aerobically and photoheterotrophically. The constitutive level of the enzyme activities are regulated according to the growth phases along with growth conditions. Thus, phbAB expression is not determinative in regulating the PB content. On the other hand, phbA-deleted cell AZI accumulated only <TEX>$10\%$</TEX> PHB of the wild-type, and an elevated dosage of phbAB in trans in R. sphaeroides resulted in a higher content of PHB, indicating that phbAB codes for the enzymes responsible for providing the main supply of subsyrate for PHB synthase. PHB formation by an alternative pathway that does not does not depend on the phbA-and phbB-coding enzymes is also proposed.