title: Purification and characterization of a mitochondrial, single-stranded-DNA-binding protein from Paracentrotus lividus eggs.
abstract: A binding protein for single-stranded DNA was purified from Paracentrotus lividus egg mitochondria to near homogeneity by chromatography on DEAE-Sephacel and single-stranded-DNA-cellulose. The protein consists of a single polypeptide of about 15 kDa. Glycerol gradient sedimentation analysis suggested that P. lividus mitochondrial single-stranded-DNA-binding protein exists as a homo-oligomer, possibly a tetramer, in solution. The protein shows a stronger preference for poly(dT) with respect to single-stranded M13, poly(dI) and poly(dC). Binding to poly(dA) takes place with much lower affinity. The binding-site size, determined by gel mobility-shift experiments with oligonucleotides of different length, is approximately 45 nucleotides. The binding to single-stranded DNA occurs with low or no cooperativity and is not influenced by ionic strength. The protein has a very high affinity for the DNA: its apparent macroscopic association constant is 2x10(9) M(-1), a value which is the highest among the mitochondrial single-stranded-DNA-binding proteins characterized to date. The lack of cooperativity and the high association constant represent distinctive features of this protein and might be related to the peculiar mechanism of sea urchin mitochondrial DNA replication.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Bari and Centro Studi sui Mitocondri e Metabolismo Energetico, CNR, Italy.
authors.forename: M
authors.initials: M
authors.lastname: Roberti
authors.forename: C
authors.initials: C
authors.lastname: Musicco
authors.forename: P
authors.initials: P
authors.lastname: Loguercio Polosa
authors.forename: M N
authors.initials: MN
authors.lastname: Gadaleta
authors.forename: E
authors.initials: E
authors.lastname: Quagliariello
authors.forename: P
authors.initials: P
authors.lastname: Cantatore
mesh_terms: D000818:Animals; D004277:DNA, Single-Stranded / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D008928:Mitochondria / Q000737:chemistry*; D010063:Ovum / Q000737:chemistry; D012617:Sea Urchins / Q000737:chemistry*
chemical_list: D004277:DNA, Single-Stranded; D004268:DNA-Binding Proteins