Lumbricus terrestris hemoglobin - The architecture of linker chains and structural variation of the central toroid

The extracellular giant hemoglobin from the earthworm Lumbricus terrestris was reconstructed at 14.9-Angstrom resolution from cryo-electron microscope images, using a new procedure for estimating parameters of the contrast tr ansfer (CTF) function. In this approach, two important CTF parameters, defo cus and amplitude contrast ratio, can be refined iteratively within the fra mework of 3D projection alignment procedure, using minimization of sign dis agreement between theoretical CTF and cross-resolution curves. The 3D cryo- EM map is in overall good agreement with the recent X-ray crystallography m ap of Royer et al (2000, Proc. Natl Acad. Sci. USA 97,7107-7111), and it re veals the local threefold arrangement of the three linker chains present wi thin each 1/12 of the complex. The 144 globin chains and 36 linker chains w ithin the complex are clearly visible, and the interdigitation of the 12 co iled-coil helical spokes forming the central toroidal piece is confirmed. B ased on these findings, two mechanisms of the dodecameric unit assembly are proposed and termed "zigzag" and "pairwise" polymerizations. However, the detection by cryo-EM of 12 additional rod-like bodies within the toroid rai ses the possibility that the architecture of the toroid is more complex tha n previously thought or that yet unknown ligands or allosteric effectors fo r this oxygen carrier are present. (C) 2001 Academic Press.