{"corpus_id":22131720,"paper_sha":"8a4c07433dc82ae8cfc5cb8754919bdf8225df8c","doi":"10.1016/S0021-9258(18)33184-3","arxiv_id":null,"pmid":6185476,"pmcid":null,"mag_id":1596622788,"dblp_id":null,"acl_id":null,"title":"Cloning of the MspI modification enzyme. The site of modification and its effects on cleavage by MspI and HpaII.","year":1983,"publication_date":"1983-01-25","venue":"Journal of Biological Chemistry","journal":{"name":"The Journal of biological chemistry","pages":"\n          1235-41\n        ","volume":"258 2"},"journal_issn":null,"journal_title":null,"publication_types":["JournalArticle"],"pubmed_pub_types":["Journal Article"],"s2_fields_of_study":["Biology","Medicine","Chemistry"],"reference_count":49,"citation_count":68,"influential_citation_count":1,"is_open_access":false,"arxiv_categories":null,"arxiv_license":null,"arxiv_journal_ref":null,"mesh_headings":[{"d":"Cloning, Molecular","mj":true,"ui":"D003001"},{"d":"DNA Restriction Enzymes","mj":false,"qs":[{"q":"genetics","mj":true,"ui":"Q000235"},{"q":"metabolism","mj":false,"ui":"Q000378"}],"ui":"D004262"},{"d":"Deoxyribonuclease HpaII","mj":false,"ui":"D019003"},{"d":"Escherichia coli","mj":false,"ui":"D004926"},{"d":"Methyltransferases","mj":false,"qs":[{"q":"metabolism","mj":false,"ui":"Q000378"}],"ui":"D008780"},{"d":"Moraxella","mj":false,"qs":[{"q":"enzymology","mj":true,"ui":"Q000201"}],"ui":"D009016"},{"d":"Plasmids","mj":false,"ui":"D010957"},{"d":"S-Adenosylmethionine","mj":false,"qs":[{"q":"metabolism","mj":false,"ui":"Q000378"}],"ui":"D012436"}],"chemicals":[{"n":"S-Adenosylmethionine","ui":"D012436","reg":"7LP2MPO46S"},{"n":"Methyltransferases","ui":"D008780","reg":"EC 2.1.1.-"},{"n":"DNA Restriction Enzymes","ui":"D004262","reg":"EC 3.1.21.-"},{"n":"Deoxyribonuclease HpaII","ui":"D019003","reg":"EC 3.1.21.-"}],"comments_corrections":null,"source_flags":5,"s2_open_access_pdf_url":null,"s2_open_access_landing_url":null,"s2_open_access_license":null,"s2_open_access_status":null,"pmc_open_access_pdf_url":null,"pmc_open_access_landing_url":null,"pmc_open_access_license":null,"pmc_open_access_status":null,"unpaywall_open_access_pdf_url":null,"unpaywall_open_access_landing_url":null,"unpaywall_open_access_license":null,"unpaywall_open_access_status":null,"abstract":"The gene for the MspI modification enzyme from Moraxella was cloned in Escherichia coli using the plasmid vector pBR322. Selection of transformants carrying the gene was based on the resistance of the modified plasmid encoding the enzyme to cleavage by MspI. Both chromosomal and plasmid DNA were modified in the selected clones. None of the clones obtained produced the cognate restriction enzyme which suggests that in this system the genes for the restriction enzyme and methylase are not closely linked. Crude cell extracts prepared from the recombinant strains, but not the host (E. coli HB101), contain an S-adenosylmethionine-dependent methyltransferase specific for the MspI recognition site, CCGG. Production of the enzyme is 3-4-fold greater in the transformants than in the original Moraxella strain. 5-Methylcytosine was identified as the product of the reaction chromatographically. The outer cytosine of the recognition sequence, *CCGG, was shown to be the site of methylation by DNA-sequencing methods. This modification blocks cleavage by both MspI and its isoschizomer HpaII. HpaII, but not MspI, is able to cleave the unmethylated strand of a hemimethylated substrate. The relevance of these results to the use of MspI and HpaII to analyze patterns of methylation in genomic DNA is discussed.","claims":[{"public_id":"cl_cefdf4786829f3e8bbb1106f9f205fef","status":"active","text":"Selection of transformants relied on resistance of the modified plasmid to cleavage by MspI.","confidence":0.95,"contributors":[{"id":1,"public_id":"12632b8b5f","public_label":"Anonymous (12632b8b5f)","roles":["extraction"],"url":"https://sah.borca.ai/u/12632b8b5f"}],"url":"https://sah.borca.ai/claims/cl_cefdf4786829f3e8bbb1106f9f205fef"},{"public_id":"cl_57bd8245ceb030057cf10d5c062687c3","status":"active","text":"The MspI modification gene from Moraxella was cloned in Escherichia coli using plasmid vector pBR322.","confidence":0.99,"contributors":[{"id":1,"public_id":"12632b8b5f","public_label":"Anonymous (12632b8b5f)","roles":["extraction"],"url":"https://sah.borca.ai/u/12632b8b5f"}],"url":"https://sah.borca.ai/claims/cl_57bd8245ceb030057cf10d5c062687c3"},{"public_id":"cl_16ec2701f74f6028c588cf020d7a1ebd","status":"active","text":"The outer cytosine of the CCGG recognition sequence is the methylation site, and this modification blocks cleavage by both MspI and HpaII; HpaII, but not MspI, can cleave the unmethylated strand of a hemimethylated substrate.","confidence":0.98,"contributors":[{"id":1,"public_id":"12632b8b5f","public_label":"Anonymous (12632b8b5f)","roles":["extraction"],"url":"https://sah.borca.ai/u/12632b8b5f"}],"url":"https://sah.borca.ai/claims/cl_16ec2701f74f6028c588cf020d7a1ebd"},{"public_id":"cl_f50f9b7411d9b57700d4ab4db7547020","status":"active","text":"The recombinant clones modified both chromosomal DNA and plasmid DNA, but none produced the cognate restriction enzyme, suggesting the restriction enzyme and methylase genes are not closely linked in this system.","confidence":0.98,"contributors":[{"id":1,"public_id":"12632b8b5f","public_label":"Anonymous (12632b8b5f)","roles":["extraction"],"url":"https://sah.borca.ai/u/12632b8b5f"}],"url":"https://sah.borca.ai/claims/cl_f50f9b7411d9b57700d4ab4db7547020"},{"public_id":"cl_563209127c6c05687a1e372e93c8fc1f","status":"active","text":"The recombinant strains contained an S-adenosylmethionine-dependent methyltransferase specific for the MspI recognition site, and enzyme production was 3-4-fold higher than in the original Moraxella strain.","confidence":0.97,"contributors":[{"id":1,"public_id":"12632b8b5f","public_label":"Anonymous (12632b8b5f)","roles":["extraction"],"url":"https://sah.borca.ai/u/12632b8b5f"}],"url":"https://sah.borca.ai/claims/cl_563209127c6c05687a1e372e93c8fc1f"}],"concepts":[{"public_id":"co_16f96e64a7c3e597e3392bd993163736","status":"active","name":"Escherichia coli","description":"The bacterial host used for cloning the MspI modification gene.","types":["host 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