title: Purification and Comparative Assay of Human Mitochondrial Single-Stranded DNA-Binding Protein.
abstract: The mitochondrial single-stranded DNA-binding protein (mtSSB) coordinates the function of replisome components at the mitochondrial replication fork. In recent years, it has been demonstrated that mtSSB stimulates the activities of DNA polymerase γ (Pol γ) and mitochondrial DNA (mtDNA) helicase in a concentration-dependent manner. Here we present a new approach to purify the human mtSSB and our standard assays to evaluate its biochemical properties, including a Gel Mobility Shift Assay (GMSA) to assess single-stranded DNA (ssDNA) binding activity, and an assay to assess SSB stimulation of Pol γ activity.
authors.affiliation: Institute of Biosciences and Medical Technology, University of Tampere, Tampere, 33014, Finland.
authors.forename: Grzegorz L
authors.initials: GL
authors.lastname: Ciesielski
authors.affiliation: Department of Biochemistry and Molecular Biology, Center for Mitochondrial Science and Medicine, Michigan State University, East Lansing, MI, 48824-1319, USA.
authors.forename: Fernando A
authors.initials: FA
authors.lastname: Rosado-Ruiz
authors.affiliation: Institute of Biosciences and Medical Technology, University of Tampere, Tampere, 33014, Finland. lskaguni@msu.edu.|Department of Biochemistry and Molecular Biology, Center for Mitochondrial Science and Medicine, Michigan State University, East Lansing, MI, 48824-1319, USA. lskaguni@msu.edu.
authors.forename: Laurie S
authors.initials: LS
authors.lastname: Kaguni
mesh_terms: D004265:DNA Helicases / Q000378:metabolism; D000074002:DNA Polymerase gamma; D004261:DNA Replication / Q000235:genetics*; D004272:DNA, Mitochondrial / Q000235:genetics*; D004277:DNA, Single-Stranded / Q000235:genetics*; D004268:DNA-Binding Proteins / Q000235:genetics*; D004259:DNA-Directed DNA Polymerase / Q000235:genetics / Q000378:metabolism*; D024202:Electrophoretic Mobility Shift Assay / Q000379:methods; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008928:Mitochondria / Q000235:genetics / Q000378:metabolism; D024101:Mitochondrial Proteins / Q000235:genetics*
chemical_list: D004272:DNA, Mitochondrial; D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins; D024101:Mitochondrial Proteins; C504091:NABP2 protein, human; D000074002:DNA Polymerase gamma; D004259:DNA-Directed DNA Polymerase; D004265:DNA Helicases
keywords: DNA polymerase γ; Human; Mitochondrial DNA replication; Mitochondrial single-stranded DNA-binding protein