X-ray crystallographic and analytical ultracentrifugation analyses of truncated and full-length yeast copper chaperones for SOD (LYS7): A dimer-dimermodel of LYS7-SOD association and copper delivery
Lt. Hall et al., X-ray crystallographic and analytical ultracentrifugation analyses of truncated and full-length yeast copper chaperones for SOD (LYS7): A dimer-dimermodel of LYS7-SOD association and copper delivery, BIOCHEM, 39(13), 2000, pp. 3611-3623
Copper-zinc superoxide dismutase (CuZnSOD) acquires its catalytic copper io
n through interaction with another polypeptide termed the copper chaperone
for SOD. Here, we combine X-ray crystallographic and analytical ultracentri
fugation methods to characterize rigorously both truncated and full-length
forms of apo-LYS7, the yeast copper chaperone for SOD. The 1.55 Angstrom cr
ystal structure of LYS7 domain 2 alone (L7D2) was determined by multiple-is
omorphous replacement (MIR) methods. The monomeric structure reveals an eig
ht-stranded Greek key beta-barrel similar to that found in yeast CuZnSOD, b
ut it is substantially elongated at one end where the loop regions of the b
eta-barrel come together to bind a calcium ion. In agreement with the cryst
al structure, sedimentation velocity experiments indicate that L7D2 is mono
meric in solution under all conditions and concentrations that were tested.
In contrast, sedimentation velocity and sedimentation equilibrium experime
nts show that full-length apo-LYS7 exists in a monomer-dimer equilibrium un
der nonreducing conditions. This equilibrium is shifted toward the dimer by
approximately 1 order of magnitude in the presence of phosphate anion. Alt
hough the basis for the specificity of the LYS7-SOD interaction as well as
the exact mechanism of copper insertion into SOD is unknown, it has been su
ggested that a monomer of LYS7 and a monomer of SOD may associate to form a
heterodimer via L7D2. The data presented here, however, taken together wit
h previously published crystallographic and analytical gel filtration data
on full-length LYS7, suggest an alternative model wherein a dimer of LYS7 i
nteracts with a dimer of yeast CuZnSOD. The advantages of the dimer-dimer m
odel over the heterodimer model are enumerated.