title: Human telomerase domain interactions capture DNA for TEN domain-dependent processive elongation.
abstract: Eukaryotic chromosome maintenance requires telomeric repeat synthesis by telomerase. It remains uncertain how telomerase domains interact to organize the active RNP and how this architecture establishes the specificity of the catalytic cycle. We combine human telomerase reconstitutions in vivo, affinity purifications, and discriminating activity assays to uncover a network of protein-protein and protein-RNA domain interactions. Notably, we find that complete single-repeat synthesis requires only a telomerase reverse transcriptase (TERT) core. Single-repeat synthesis does not require the TERT N-terminal (TEN) domain, but RNA-dependent positioning of the TEN domain captures substrate and allows repeat synthesis processivity. A TEN domain physically separate from the TERT core can capture even a minimal template-paired DNA substrate, with substrate association enhanced by the presence of a 5' single-stranded extension. Our results provide insights into active enzyme architecture, explain biological variations of the catalytic cycle, and predict altered activities for TERT proteins of some eukaryotes.
authors.affiliation: Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720-3200, USA.
authors.forename: Aaron R
authors.initials: AR
authors.lastname: Robart
authors.forename: Kathleen
authors.initials: K
authors.lastname: Collins
mesh_terms: D001665:Binding Sites; D055162:Biocatalysis; D002460:Cell Line; D004247:DNA / Q000235:genetics / Q000378:metabolism*; D004277:DNA, Single-Stranded / Q000235:genetics / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012313:RNA / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D019098:Telomerase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D016615:Telomere / Q000235:genetics / Q000378:metabolism
chemical_list: D004277:DNA, Single-Stranded; C422952:telomerase RNA; D012313:RNA; D004247:DNA; C509186:TERT protein, human; D019098:Telomerase