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Modification-dependent restriction endonuclease, MspJI, flips 5-methylcytosine out of the DNA helix
MspJI belongs to a family of restriction enzymes that cleave DNA containing 5-methylcytosine (5mC) or 5-hydroxymethylcytosine (5hmC). MspJI is specific for the sequence 5(h)mC-N-N-G or A and cleaves with some variability 9/13 nucleotides downstream. Earlier, we reported the crystal structure of MspJ...
Autores principales: | , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
Oxford University Press
2014
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4231741/ https://www.ncbi.nlm.nih.gov/pubmed/25262349 http://dx.doi.org/10.1093/nar/gku871 |
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author | Horton, John R. Wang, Hua Mabuchi, Megumu Yamada Zhang, Xing Roberts, Richard J. Zheng, Yu Wilson, Geoffrey G. Cheng, Xiaodong |
author_facet | Horton, John R. Wang, Hua Mabuchi, Megumu Yamada Zhang, Xing Roberts, Richard J. Zheng, Yu Wilson, Geoffrey G. Cheng, Xiaodong |
author_sort | Horton, John R. |
collection | PubMed |
description | MspJI belongs to a family of restriction enzymes that cleave DNA containing 5-methylcytosine (5mC) or 5-hydroxymethylcytosine (5hmC). MspJI is specific for the sequence 5(h)mC-N-N-G or A and cleaves with some variability 9/13 nucleotides downstream. Earlier, we reported the crystal structure of MspJI without DNA and proposed how it might recognize this sequence and catalyze cleavage. Here we report its co-crystal structure with a 27-base pair oligonucleotide containing 5mC. This structure confirms that MspJI acts as a homotetramer and that the modified cytosine is flipped from the DNA helix into an SRA-like-binding pocket. We expected the structure to reveal two DNA molecules bound specifically to the tetramer and engaged with the enzyme's two DNA-cleavage sites. A coincidence of crystal packing precluded this organization, however. We found that each DNA molecule interacted with two adjacent tetramers, binding one specifically and the other non-specifically. The latter interaction, which prevented cleavage-site engagement, also involved base flipping and might represent the sequence-interrogation phase that precedes specific recognition. MspJI is unusual in that DNA molecules are recognized and cleaved by different subunits. Such interchange of function might explain how other complex multimeric restriction enzymes act. |
format | Online Article Text |
id | pubmed-4231741 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2014 |
publisher | Oxford University Press |
record_format | MEDLINE/PubMed |
spelling | pubmed-42317412014-11-21 Modification-dependent restriction endonuclease, MspJI, flips 5-methylcytosine out of the DNA helix Horton, John R. Wang, Hua Mabuchi, Megumu Yamada Zhang, Xing Roberts, Richard J. Zheng, Yu Wilson, Geoffrey G. Cheng, Xiaodong Nucleic Acids Res Nucleic Acid Enzymes MspJI belongs to a family of restriction enzymes that cleave DNA containing 5-methylcytosine (5mC) or 5-hydroxymethylcytosine (5hmC). MspJI is specific for the sequence 5(h)mC-N-N-G or A and cleaves with some variability 9/13 nucleotides downstream. Earlier, we reported the crystal structure of MspJI without DNA and proposed how it might recognize this sequence and catalyze cleavage. Here we report its co-crystal structure with a 27-base pair oligonucleotide containing 5mC. This structure confirms that MspJI acts as a homotetramer and that the modified cytosine is flipped from the DNA helix into an SRA-like-binding pocket. We expected the structure to reveal two DNA molecules bound specifically to the tetramer and engaged with the enzyme's two DNA-cleavage sites. A coincidence of crystal packing precluded this organization, however. We found that each DNA molecule interacted with two adjacent tetramers, binding one specifically and the other non-specifically. The latter interaction, which prevented cleavage-site engagement, also involved base flipping and might represent the sequence-interrogation phase that precedes specific recognition. MspJI is unusual in that DNA molecules are recognized and cleaved by different subunits. Such interchange of function might explain how other complex multimeric restriction enzymes act. Oxford University Press 2014-10-29 2014-09-27 /pmc/articles/PMC4231741/ /pubmed/25262349 http://dx.doi.org/10.1093/nar/gku871 Text en © The Author(s) 2014. Published by Oxford University Press on behalf of Nucleic Acids Research. http://creativecommons.org/licenses/by-nc/4.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com |
spellingShingle | Nucleic Acid Enzymes Horton, John R. Wang, Hua Mabuchi, Megumu Yamada Zhang, Xing Roberts, Richard J. Zheng, Yu Wilson, Geoffrey G. Cheng, Xiaodong Modification-dependent restriction endonuclease, MspJI, flips 5-methylcytosine out of the DNA helix |
title | Modification-dependent restriction endonuclease, MspJI, flips 5-methylcytosine out of the DNA helix |
title_full | Modification-dependent restriction endonuclease, MspJI, flips 5-methylcytosine out of the DNA helix |
title_fullStr | Modification-dependent restriction endonuclease, MspJI, flips 5-methylcytosine out of the DNA helix |
title_full_unstemmed | Modification-dependent restriction endonuclease, MspJI, flips 5-methylcytosine out of the DNA helix |
title_short | Modification-dependent restriction endonuclease, MspJI, flips 5-methylcytosine out of the DNA helix |
title_sort | modification-dependent restriction endonuclease, mspji, flips 5-methylcytosine out of the dna helix |
topic | Nucleic Acid Enzymes |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4231741/ https://www.ncbi.nlm.nih.gov/pubmed/25262349 http://dx.doi.org/10.1093/nar/gku871 |
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