Structure of the TnsB transposase-DNA complex of type V-K CRISPR-associated transposon
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CRISPR-associated transposons (CASTs) are mobile genetic elements that co-opted CRISPR-Cas systems for RNA-guided transposition. Here we present the 2.4 Å cryo-EM structure of the Scytonema hofmannii (sh) TnsB transposase from Type V-K CAST, bound to the strand transfer DNA. The strand transfer complex displays an intertwined pseudo-symmetrical architecture. Two protomers involved in strand transfer display a catalytically competent active site composed by DDE residues, while other two, which play a key structural role, show active sites where the catalytic residues are not properly positioned for phosphodiester hydrolysis. Transposon end recognition is accomplished by the NTD1/2 helical domains. A singular in trans association of NTD1 domains of the catalytically competent subunits with the inactive DDE domains reinforces the assembly. Collectively, the structural features suggest that catalysis is coupled to protein-DNA assembly to secure proper DNA integration. DNA binding residue mutants reveal that lack of specificity decreases activity, but it could increase transposition in some cases. Our structure sheds light on the strand transfer reaction of DDE transposases and offers new insights into CAST transposition.
| Originalsprog | Engelsk |
|---|---|
| Artikelnummer | 5792 |
| Tidsskrift | Nature Communications |
| Vol/bind | 13 |
| Antal sider | 14 |
| ISSN | 2041-1723 |
| DOI | |
| Status | Udgivet - 2022 |
Bibliografisk note
Funding Information:
We thank the Danish Cryo-EM National Facility in CFIM at the University of Copenhagen and specially Tillmann Pape for support during cryo-EM data collection. We also thank the Protein Expression Unit at CPR for assistance in protein expression and purification. F.T.C. is a member of the Copenhagen Bioscience PhD programme (NNF19SA0035440). G.M. is part of the Novo Nordisk Foundation Center for Protein Research (CPR), which is supported financially by the Novo Nordisk Foundation (grant NNF14CC0001). This work was also supported by grant NNF0024386, grant NNF17SA0030214 and Distinguished Investigator (NNF18OC0055061) grants to G.M., who is a member of the Integrative Structural Biology Cluster (ISBUC) at the University of Copenhagen.
Funding Information:
We thank the Danish Cryo-EM National Facility in CFIM at the University of Copenhagen and specially Tillmann Pape for support during cryo-EM data collection. We also thank the Protein Expression Unit at CPR for assistance in protein expression and purification. F.T.C. is a member of the Copenhagen Bioscience PhD programme (NNF19SA0035440). G.M. is part of the Novo Nordisk Foundation Center for Protein Research (CPR), which is supported financially by the Novo Nordisk Foundation (grant NNF14CC0001). This work was also supported by grant NNF0024386, grant NNF17SA0030214 and Distinguished Investigator (NNF18OC0055061) grants to G.M., who is a member of the Integrative Structural Biology Cluster (ISBUC) at the University of Copenhagen.
Publisher Copyright:
© 2022, The Author(s).
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