טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentJacob Eyal
SubjectCharacterization of the Interactions between the Protein
Translin and Single Stranded Guanine Rich DNA
Sequences
DepartmentDepartment of Biology
Supervisor Professor Haim Manor


Abstract

We have previously identified in human fibroblasts a multisubunit protein (designated PGB) that specifically binds single-stranded G-rich microsatellite DNA sequences. PGB was later found to be identical or closely related to Translin, an octameric protein that binds single-stranded DNA consisting of sequences flanking chromosomal translocations. Here we report that recombinant Translin binds single-stranded d(GT)n microsatellites and G-strand telomeric repeats, d(TTAGGG)n, with higher affinities (Kdis@2 nM and Kdis@12.5 nM, respectively) than the affinity with which it binds translocation consensus sequences (Kdis@23 nM). A detailed study revealed that (i) the minimal length of d(GT)n and d(TTAGGG)n oligonucleotides that a Translin octamer binds is 11 nucleotides. (ii) While oligos of ≤30 nucleotides bind a single Translin octamer, d(GT)27 and d(TTAGGG)9 bind two octamers with negative cooperativity. (iii) Formation of quadruplex structures alters the binding patterns of d(TTAGGG)n repeats to Translin. (iv) Binding to DNA induces a conformational change in Translin that persists, at least temporarily, after dissociation of the DNA. (v) Translin binds with high affinity (Kdis@1.5 nM) 3’ and 5’ d(GT)n overhangs, but does not bind single-stranded d(GT)n sequences embedded within double-stranded DNA. Based on these data, we propose that Translin might be involved in recombination at d(GT)n.d(AC)n microsatellites and in telomere maintenance.