Structural dissection of sequence recognition and catalytic mechanism of human LINE-1 endonuclease
| Autor: | Max Totrov, Denis N. Kazyulkin, Lioubov G. Korotchkina, Sergey Korolev, Andrei V. Gudkov, Ian J. Miller |
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| Jazyk: | angličtina |
| Rok vydání: | 2021 |
| Předmět: |
Models
Molecular Protein Conformation alpha-Helical AcademicSubjects/SCI00010 Genetic Vectors Gene Expression Retrotransposon Computational biology Crystallography X-Ray Genomic Instability Substrate Specificity Endonuclease chemistry.chemical_compound Scissile bond Structural Biology Genetics Consensus sequence DNA-(Apurinic or Apyrimidinic Site) Lyase Escherichia coli Deoxyribonuclease I Humans AP site Protein Interaction Domains and Motifs Cloning Molecular DNA Cleavage Binding Sites biology Base Sequence Genome Human DNA Recombinant Proteins chemistry biology.protein Nucleic Acid Conformation Thermodynamics Human genome Protein Conformation beta-Strand Primer (molecular biology) Protein Binding |
| Zdroj: | Nucleic Acids Research |
| ISSN: | 1362-4962 0305-1048 |
| Popis: | Long interspersed nuclear element-1 (L1) is an autonomous non-LTR retrotransposon comprising ∼20% of the human genome. L1 self-propagation causes genomic instability and is strongly associated with aging, cancer and other diseases. The endonuclease domain of L1’s ORFp2 protein (L1-EN) initiates de novo L1 integration by nicking the consensus sequence 5′-TTTTT/AA-3′. In contrast, related nucleases including structurally conserved apurinic/apyrimidinic endonuclease 1 (APE1) are non-sequence specific. To investigate mechanisms underlying sequence recognition and catalysis by L1-EN, we solved crystal structures of L1-EN complexed with DNA substrates. This showed that conformational properties of the preferred sequence drive L1-EN’s sequence-specificity and catalysis. Unlike APE1, L1-EN does not bend the DNA helix, but rather causes ‘compression’ near the cleavage site. This provides multiple advantages for L1-EN’s role in retrotransposition including facilitating use of the nicked poly-T DNA strand as a primer for reverse transcription. We also observed two alternative conformations of the scissile bond phosphate, which allowed us to model distinct conformations for a nucleophilic attack and a transition state that are likely applicable to the entire family of nucleases. This work adds to our mechanistic understanding of L1-EN and related nucleases and should facilitate development of L1-EN inhibitors as potential anticancer and antiaging therapeutics. Graphical Abstract Graphical AbstractMechanism of DNA sequence recognition and cleavage by L1-EN endonuclease domain during initiation of the LINE-1 retrotransposon target-primed reverse transcription. |
| Databáze: | OpenAIRE |
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