title: L1 retrotransposition requires rapid ORF1p oligomerization, a novel coiled coil-dependent property conserved despite extensive remodeling.
abstract: Detailed mechanistic understanding of L1 retrotransposition is sparse, particularly with respect to ORF1p, a coiled coil-mediated homotrimeric nucleic acid chaperone that can form tightly packed oligomers on nucleic acids. Although the coiled coil motif is highly conserved, it is uniquely susceptible to evolutionary change. Here we studied three ORF1 proteins: a modern human one (111p), its resuscitated primate ancestor (555p) and a mosaic modern protein (151p) wherein 9 of the 30 coiled coil substitutions retain their ancestral state. While 111p and 555p equally supported retrotransposition, 151p was inactive. Nonetheless, they were fully active in bulk assays of nucleic acid interactions including chaperone activity. However, single molecule assays showed that 151p trimers form stably bound oligomers on ssDNA at <1/10th the rate of the active proteins, revealing that oligomerization rate is a novel critical parameter of ORF1p activity in retrotransposition conserved for at least the last 25 Myr of primate evolution.
authors.affiliation: Northeastern University, Department of Physics, Boston, MA 02115, USA.
authors.forename: M Nabuan
authors.initials: MN
authors.lastname: Naufer
authors.affiliation: Laboratory of Cellular and Molecular Biology, NIDDK, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Kathryn E
authors.initials: KE
authors.lastname: Callahan
authors.affiliation: Laboratory of Cellular and Molecular Biology, NIDDK, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Pamela R
authors.initials: PR
authors.lastname: Cook
authors.affiliation: Laboratory of Cellular and Molecular Biology, NIDDK, National Institutes of Health, Bethesda, MD 20892, USA.
authors.forename: Cesar E
authors.initials: CE
authors.lastname: Perez-Gonzalez
authors.affiliation: Northeastern University, Department of Physics, Boston, MA 02115, USA mark@neu.edu.
authors.forename: Mark C
authors.initials: MC
authors.lastname: Williams
authors.affiliation: Laboratory of Cellular and Molecular Biology, NIDDK, National Institutes of Health, Bethesda, MD 20892, USA avf@helix.nih.gov.
authors.forename: Anthony V
authors.initials: AV
authors.lastname: Furano
mesh_terms: D004277:DNA, Single-Stranded; D006801:Humans; D007700:Kinetics; D020084:Long Interspersed Nucleotide Elements / Q000235:genetics*; D018832:Molecular Chaperones / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D009841:Oligonucleotides / Q000235:genetics / Q000378:metabolism; D011485:Protein Binding; D055503:Protein Multimerization; D011506:Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D004277:DNA, Single-Stranded; D018832:Molecular Chaperones; C531245:ORF1 protein, human; D009841:Oligonucleotides; D011506:Proteins