Association of a protein structure of probable membrane derivation with HeLa cell mitochondrial DNA near its origin of replication.

Abstract

Almost all (about 95%) of the mitochondrial DNA molecules released by Triton X-100 lysis of HeLa [human cervical carcinoma] cell mitochondria in the presence of 0.15 M salt are associated with a single protein-containing structure varying in appearance between a 10-20 nm knob and a 100-500 nm membrane-like patch. Analysis by high resolution EM and by polyacrylamide gel electrophoresis after cleavage of mitochondrial DNA with the endonucleases EcoRI, HindIII and Hpa II has shown that the protein structure is attached to the DNA in the region of the D-loop, and probably near the origin of mitochondrial DNA replication. HeLa cell mitochondrial DNA is probably attached in vivo to the inner mitochondrial membrane at or near the origin of replication. A membrane fragment of variable size may remain associated with the DNA during the isolation. After sodium dodecyl sulfate extraction of mitochondrial DNA, a small 5-10 nm protein is found at the same site on a fraction of the mitochondrial DNA molecules.