Crystal structures of the regulatory subunit of Thr-sensitive aspartate kinase from Thermus thermophilus

Abstract

Crystal structures of the regulatory subunit of Thr-sensitive aspartate kinase (AK; EC 2.7.2.4) from Thermus thermophilus (TtAK) were determined at 2.15 a in the Thr-bound form (TtAK-Thr) and at 2.98 a in the Thr-free form (TtAK-free). Although both forms are crystallized as dimers, the contact surface area of the dimer interface in TtAK-free (3200 a2) is smaller than that of TtAK-Thr (3890 a2). Sedimentation equilibrium analyzed by ultracentrifugation revealed that TtAK is present in equilibrium between a monomer and dimer, and that Thr binding shifts the equilibrium to dimer formation. In the absence of Thr, an outward shift of -strands near the Thr-binding site (site 1) and a concomitant loss of the electron density of the loop region between 3 and 4 near the Thr-binding site are observed. The mechanism of regulation by Thr is discussed on the basis of the crystal structures. TtAK has higher thermostability than the regulatory subunit of Corynebacterium glutamicum AK, with a difference in denaturation temperature (Tm) of 40 °C. Comparison of the crystal structures of TtAK and the regulatory subunit of C. glutamicum AK showed that the well-packed hydrophobic core and high Pro content in loops contribute to the high thermostability of TtAK. © 2009 FEBS.