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Durum wheat metallothionein (dMT), a plant type 1 metallothionein, with a long “hinge” region between metal coordinating cysteine clusters, is efficient cadmium (Cd) chelator. In this work, biophysical features of purified recombinant holo-dMT, its demetallated form (apo-dMT) and the reconstituted Cd5-dMT are compared to obtain insight into the structure and metal binding features of this protein. Results show that the purified holo-dMT is polydisperse and has 5.3±0.5 Cd2+ ions per molecule. Demetallation followed by size exclusion chromatography yields homogeneous apo-dMT which can be reconstituted with Cd2+. Synchrotron small angle X-ray scattering (SAXS) demonstrates that apo-dMT, at pH 2.0, is flexible and extended in solution. According to UV-vis, CD and native-PAGE data conformation of apo-dMT is sensitive to pH changes in the range 2.0 to 8.0. Reconstitution of the apo-protein at pH 8.0, with Cd2+ appears to take place in two phases during which first the monomer is folded to accommodate 5 Cd2+ ions and then reorganization into oligomeric forms allows incorporation of further metal ions. SAXS data indicate that holo-dMT has limited flexibility in structure, but its conformation is significantly more compact than that of apo-dMT. Results of UV-vis and circular dichroism spectroscopy show that the in vitro folded protein is structurally different from the purified holo-dMT with the same number of Cd2+ ions.This work was supported by Sabanci University Internal Research Fund Project No. IACF08-00514 and by the bilateral program TUBITAK-Julich Research Center projects TBAG-U-155 and TBAG-U-157.

Calculation of the size of the power stroke of the myosin motor in contracting muscle requires knowledge of the compliance of the myofilaments. Current estimates of actin compliance vary significantly introducing uncertainty in the mechanical parameters of the motor. Using x-ray diffraction on small bundles of permeabilized fibers from rabbit muscle we show that strong binding of myosin heads changes directly the actin helix. The spacing of the 2.73-nm meridional x-ray reflection increased by 0.22% when relaxed fibers were put into low-tension rigor (