Asp295 Stabilizes the Active-Site Loop Structure of Pyruvate Dehydrogenase, Facilitating Phosphorylation of Ser292 by Pyruvate Dehydrogenase-Kinase

Figure 6

Quantitative analysis of interactions between AtPDC E1 and AtPDK. Quantitative B2H analysis was used to study interaction between WT AtPDC E1α, the D295A, and D295N mutants and AtPDK. The AtPDK sequence was cloned into the Stratagene B2H vector pBT, while the E1α sequences were cloned into pTRG. After selection, protein interactions were quantified using the β-galactosidase reporter. The value for 100% of WT activity was 3 μmol min⁻¹ 1.0 A600⁻¹. Data are the mean ± SEM from two separate experiments, each assay conducted in triplicate.