3-DIMENSIONAL RECONSTRUCTION OF A COMPLEX OF HUMAN ALPHA(2)-MACROGLOBULIN WITH MONOMALEIMIDO NANOGOLD (AU(1.4NM)) EMBEDDED IN ICE

Cysteine 949 and glutamine 952 are known to be part of the thiol ester site of each of the four subunits of human α2-macroglobulin (α2M). The hydrolysis of this thiol ester bound to methylamine results in the incorporation of the amine and liberation of a free sulfhydryl group that can be specifically labeled. Therefore, a high-resolution marker specific for the sulfhydryl groups, the monomaleimido Nanogold (Au1.4nm) cluster was used to bind this amino acid. After cryoelectron microscopy, a three-dimensional reconstruction of the α2M-Nanogold conjugates (α2M-Au1.4nm) was achieved, revealing the internal location of the thiol ester sites in the transformed α2M molecules. From this study we propose three possible locations for the cysteine 949. © 1992.