Mitochondrial localization and oligomeric structure of HClpP, the human homologue of E. coli ClpP.
A bacterially expressed recombinant HClpP protein, the human homologue of Escherichia coli ClpP protease, was used to obtain specific polyclonal antibodies. Those antibodies identify a 26 kDa polypeptide in mitochondrial subcellular fractions of rat and human liver. Immunofluorescence and electron microscopic studies demonstrate that the mammalian homologue of ClpP is located in the mitochondrial matrix with a tendency to be found in association with the inner mitochondrial membrane. An HClpP recombinant protein with a truncated NH2terminus (missing the first 58 amino acid residues) shows a molecular mass of 26 kDa under denaturing conditions. This N-truncated HClpP recombinant protein shows a native molecular mass of 340 kDa that is identical with the native molecular mass of the partially purified protein from rat liver mitochondria. Electron microscopy shows that the N-truncated recombinant HClpP has a ring shape with seven identical morphological units in the periphery, exhibiting a 7-fold symmetry. The native molecular mass and the electron microscopic studies suggest that mitochondrial ClpP is composed of two heptameric rings with 7-fold symmetry, similar to E. coli ClpP.
de Sagarra
M R
MR
Departamento de Bioquímica e Instituto de Investigaciones Biomédicas "Alberto Sols". UAM-CSIC, Facultad de Medicina UAM, Madrid, 28029, Spain.
Mayo
I
I
Marco
S
S
Rodríguez-Vilariño
S
S
Oliva
J
J
Carrascosa
J L
JL
Casta ñ
J G
JG
D000251:Adenosine Triphosphatases / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D002460:Cell Line; D002850:Chromatography, Gel; D049071:Endopeptidase Clp; D004926:Escherichia coli / Q000737:chemistry / Q000235:genetics; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism; D016253:Microscopy, Immunoelectron; D008930:Mitochondria, Liver / Q000737:chemistry* / Q000378:metabolism / Q000648:ultrastructure; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011485:Protein Binding; D011487:Protein Conformation; D051381:Rats; D011994:Recombinant Proteins / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D017384:Sequence Deletion; D012697:Serine Endopeptidases / Q000032:analysis / Q000737:chemistry* / Q000378:metabolism*
D000906:Antibodies; D011994:Recombinant Proteins; D012697:Serine Endopeptidases; D049071:Endopeptidase Clp; D000251:Adenosine Triphosphatases