Topology of the morphological domains of the chaperonin GroEL visualized by immuno-electron microscopy.
Electron microscopy of the tetradecameric double-ring complex of GroEL reveals a four-layered structure, indicating that the 58 kDa subunits are composed of two major morphological domains. We have used immuno-electron microscopy to assign these domains to the corresponding segments of the GroEL sequence. Upon chemical modification of GroEL with N-ethylmaleimide, protease treatment in the presence of ATP or ADP generates GroEL fragments of 15 kDa (N15; residues 1-141) and 40 kDa (C40; residues 153-531). As visualized by scanning transmission electron microscopy, affinity-purified antibodies directed against C40 recognize the outer layers, whereas antibodies against N15 interact with the equatorial portions of the GroEL double-ring. Thus, the two major domains of the subunits in the chaperonin complex are arranged in the order C40-N15:N15-C40. The single-ring chaperonin co-factor GroES interacts with the C40 domain while the ATP-binding site of GroEL is probably close to the junction between N15 and C40.
Cellular Biochemistry & Biophysics Program, Memorial Sloan-Kettering Cancer Center, New York, New York 10021.
J
J
Martin
K N
KN
Goldie
A
A
Engel
F U
FU
Hartl
D000906:Antibodies / Q000302:isolation & purification; D018834:Chaperonin 60 / Q000737:chemistry / Q000648:ultrastructure*; D046911:Macromolecular Substances; D017348:Microscopy, Electron, Scanning Transmission; D016253:Microscopy, Immunoelectron / Q000379:methods; D008970:Molecular Weight; D010446:Peptide Fragments / Q000737:chemistry / Q000302:isolation & purification
D000906:Antibodies; D018834:Chaperonin 60; D046911:Macromolecular Substances; D010446:Peptide Fragments