Ca2+-loaded spherulin 3a from Physarum polycephalum adopts the prototype gamma-crystallin fold in aqueous solution

Spherulin 3a is the most abundantly expressed cytosolic protein in spherulating plasmodia of the slime mold Physarum polycephalum. High yields of unlabeled, uniformly N-15 and uniformly C-13/N-15-labeled recombinant spherulin 3a from Escherichia coli could be produced by a simple protocol described here. The three-dimensional solution structure of Ca2+-loaded spherulin 3a was determined by home-and heteronuclear NMR spec troscopy. The structure of monomeric spherulin 3a consists of two pleated beta-sheets plus a short alpha-helix arranged into the gamma-crystallin fold. The beta-sheets comprise two intertwined Greek-key motifs. An additional N-terminal beta-strand is unique to spherulin 3a. Complexation of calcium ions greatly enhances overall conformational stability of the protein. The average atomic root-mean-square deviations (r.m.s.d.) for heavy atoms in beta-strands were 0.34(+/-0.16) Angstrom for the backbone atoms and 0.73(+/-0.40) Angstrom for all atoms. The corresponding r.m.s.d. values for heavy atoms in the whole protein were 0.62(+/-0.42) Angstrom for the backbone atoms and 0.99(+/-0.65) A for all atoms. We show the structural relationship between spherulin 3a, a myxomycete dormancy protein, and crystallins from the vertebrate eye lens. Since spherulin 3a has a structure corresponding to one domain of bovine gamma B(II)-crystallin, it represents a hypothetical ancestral gamma-crystallin precursor structure. (C) 1997 Academic Press Limited.