Journal: Life Science Alliance
Article Title: Defective nucleotide-dependent assembly and membrane fusion in Mfn2 CMT2A variants improved by Bax
Figure Lengend Snippet: Mitochondria were isolated from a clonal population of Mfn1-null cells expressing Mfn1 WT -eGFP at endogenous levels (Mfn1-eGFP) or a clonal population of Mfn2-null cells expressing Mfn2-FLAG, or a clonal population of Mfn1-null cells expressing Mfn1-FLAG. Mitochondria were combined in the indicated combinations and incubated with BeF 3 in the presence or absence of GDP. After lysis, immunoprecipitation was performed with α-FLAG magnetic beads. Proteins eluted from the beads were subjected to SDS–PAGE and immunoblotting with α-Mfn1, α-FLAG, α-VDAC, or α-Hsp60, as indicated. Black arrow indicates endogenous Mfn1, white arrowhead indicates Mfn1-Egfp, black arrowhead indicates Mfn2-FLAG, and line indicates Mfn1-FLAG. Input represents 3% of the input and elution represents 37.5% of the immunoprecipitated protein. The molecular weight markers are shown in kD on the left.
Article Snippet: MEF cells (Mfn wild-type and Mfn2-null) were purchased from American Type Culture Collection.
Techniques: Isolation, Expressing, Incubation, Lysis, Immunoprecipitation, Magnetic Beads, SDS Page, Western Blot, Molecular Weight