The binding in vitro of the intermediate filament protein vimentin to synthetic oligonucleotides containing telomere sequences.
The ability of the intermediate filament subunit protein vimentin to bind synthetic oligonucleotide telomere models containing repeat sequences from Oxytricha (T4G4), Saccharomyces (TGTGTG3), or Tetrahymena (T2G4) was investigated in vitro with a filter binding assay and a gel overlay assay. At low ionic strength, vimentin bound these oligonucleotides with high affinity. At higher ionic strength, the vimentin-oligonucleotide complex was less stable, such that approximately 30% of the initial binding remained at 150 mM KCl. One mole of vimentin tetramer bound approximately 1 mol of telomere oligonucleotide. Vimentin bound well oligonucleotides containing either a random duplex or random 3'-overhang, but showed a reduced affinity for a blunt-ended oligonucleotide. A control random sequence oligonucleotide was not bound by vimentin. The oligonucleotide-binding site of vimentin was shown to be localized in the non-alpha-helical N-terminal domain by assays employing purified proteolytic fragments of vimentin. Preliminary results in the gel overlay assay show that other members of the intermediate filament family, nuclear lamins A-C, all bind the synthetic oligonucleotide containing the telomere repeat sequence of Oxytricha.
Shoeman
R L
RL
Max-Planck-Institut für Zellbiologie, Ladenburg bei Heidelberg, Federal Republic of Germany.
Wadle
S
S
Scherbarth
A
A
Traub
P
P
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D015252:Deoxyribonucleases, Type II Site-Specific / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D009841:Oligonucleotides / Q000378:metabolism*; D012091:Repetitive Sequences, Nucleic Acid; D012440:Saccharomyces; D013329:Structure-Activity Relationship; D013768:Tetrahymena; D014746:Vimentin / Q000378:metabolism*
D009841:Oligonucleotides; D014746:Vimentin; D015252:Deoxyribonucleases, Type II Site-Specific; C024403:GGCC-specific type II deoxyribonucleases