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Time course of CoA release from HBCoA at 30 °C catalyzed by Strep2-tagged synthases purified from R. eutropha at 24 h and 4 h TSB growth and from E. coli. The reaction mixtures for Strep2-PhaCRe24h (□), Strep2-PhaCEc (●), and C319A Strep2-PhaCRe (×) contain 60 nM enzyme and 750 μM HBCoA. The reaction mixture for Strep2-PhaCRe4h (▲) contains 60 nM enzyme and 600 μM HBCoA. HBCoA concentrations used in the assays are well above the Km (100 μM) (17).
