Salt-Induced Assembly Kinetics of Archaeoglobus Fulgidus Ferritin Monitored By Time-Resolved Small-Angle X-Ray Scattering

Ferritin is a protein cage composed of 24 identical subunits with octahedral 4-3-2 symmetry. Archaoglobus fulgidus ferritin (AfFtnWT) has a unique “open structure” shifting from 4-fold symmetry to tetrahedral 2-3 symmetry forming 4 triangular pores with 52Å edges in the shell. It has been shown that the replacements of Lys-150 and Arg-151 to alanines dramatically change the assembly of AfFtn. The mutant K150A/R151A (AfFtnAA) assembled into a conventional octahedral cage, in which Ala-150 and Arg-151 were located at 4-fold symmetry axis. Furthermore, AfFtnWT and AfFtnAA exist as subunit dimers having native-like secondary and tertiary structures at low salt concentrations. They can reversibly reassemble into respective 24mers when salt concentration is increased. In this study, the assembly reactions of AfFtnWT and AfFtnAA were induced by NaCl-concentration jump and monitored by time-resolved small-angle x-ray scattering (TR-SAXS). The results indicated that the assembly rate of AfFtnAA is 25 times faster than that of AfFtnWT of 445mM NaCl. Although the detailed analysis of the assembly reaction was hampered by the unknown large aggregate formation at higher protein concentrations under the low salt conditions, the kinetic curve of AfFtnAA was similar to that of Escherichia coli ferritin (EcFtnA), indicating that AfFtnAA assembles into 24mer via the mechanism similar to that previously elucidated for EcFtnA.