Structural and Functional Characterization of Rv2966c Protein Reveals an RsmD-like Methyltransferase from Mycobacterium tuberculosis and the Role of Its N-terminal Domain in Target Recognition
Nine of ten methylated nucleotides of Escherichia coli 16 S rRNA are conserved in Mycobacterium tuberculosis. All the 10 different methyltransferases are known in E. coli, whereas only TlyA and GidB have been identified in mycobacteria. Here we have identified Rv2966c of M. tuberculosis as an orthol...
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| Veröffentlicht in: | The Journal of biological chemistry 2011-06, Vol.286 (22), p.19652-19661 |
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| container_title | The Journal of biological chemistry |
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| creator | Kumar, Atul Saigal, Kashyap Malhotra, Ketan Sinha, Krishna Murari Taneja, Bhupesh |
| description | Nine of ten methylated nucleotides of Escherichia coli 16 S rRNA are conserved in Mycobacterium tuberculosis. All the 10 different methyltransferases are known in E. coli, whereas only TlyA and GidB have been identified in mycobacteria. Here we have identified Rv2966c of M. tuberculosis as an ortholog of RsmD protein of E. coli. We have shown that rv2966c can complement rsmD-deleted E. coli cells. Recombinant Rv2966c can use 30 S ribosomes purified from rsmD-deleted E. coli as substrate and methylate G966 of 16 S rRNA in vitro. Structure determination of the protein shows the protein to be a two-domain structure with a short hairpin domain at the N terminus and a C-terminal domain with the S-adenosylmethionine-MT-fold. We show that the N-terminal hairpin is a minimalist functional domain that helps Rv2966c in target recognition. Deletion of the N-terminal domain prevents binding to nucleic acid substrates, and the truncated protein fails to carry out the m2G966 methylation on 16 S rRNA. The N-terminal domain also binds DNA efficiently, a property that may be utilized under specific conditions of cellular growth. |
| doi_str_mv | 10.1074/jbc.M110.200428 |
| format | Article |
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All the 10 different methyltransferases are known in E. coli, whereas only TlyA and GidB have been identified in mycobacteria. Here we have identified Rv2966c of M. tuberculosis as an ortholog of RsmD protein of E. coli. We have shown that rv2966c can complement rsmD-deleted E. coli cells. Recombinant Rv2966c can use 30 S ribosomes purified from rsmD-deleted E. coli as substrate and methylate G966 of 16 S rRNA in vitro. Structure determination of the protein shows the protein to be a two-domain structure with a short hairpin domain at the N terminus and a C-terminal domain with the S-adenosylmethionine-MT-fold. We show that the N-terminal hairpin is a minimalist functional domain that helps Rv2966c in target recognition. Deletion of the N-terminal domain prevents binding to nucleic acid substrates, and the truncated protein fails to carry out the m2G966 methylation on 16 S rRNA. The N-terminal domain also binds DNA efficiently, a property that may be utilized under specific conditions of cellular growth.</description><identifier>ISSN: 0021-9258</identifier><identifier>EISSN: 1083-351X</identifier><identifier>DOI: 10.1074/jbc.M110.200428</identifier><identifier>PMID: 21474448</identifier><language>eng</language><publisher>United States: Elsevier Inc</publisher><subject>Bacterial Proteins - chemistry ; Bacterial Proteins - genetics ; Bacterial Proteins - metabolism ; Crystal Structure ; DNA-binding Protein ; Histones - chemistry ; Histones - genetics ; Histones - metabolism ; Methylation ; Mycobacterium tuberculosis - enzymology ; Mycobacterium tuberculosis - genetics ; Protein Structure ; Protein Structure and Folding ; Protein Structure, Tertiary ; Ribosomal RNA (rRNA) ; RNA Methylation ; RNA Methyltransferase ; RNA, Bacterial - chemistry ; RNA, Bacterial - genetics ; RNA, Bacterial - metabolism ; RNA, Ribosomal, 16S - chemistry ; RNA, Ribosomal, 16S - genetics ; RNA, Ribosomal, 16S - metabolism ; RsmD ; Rv2966c ; S-Adenosylmethionine (SAM) ; SAM-MT-fold ; Structure-Activity Relationship ; tRNA Methyltransferases - chemistry ; tRNA Methyltransferases - genetics ; tRNA Methyltransferases - metabolism</subject><ispartof>The Journal of biological chemistry, 2011-06, Vol.286 (22), p.19652-19661</ispartof><rights>2011 © 2011 ASBMB. Currently published by Elsevier Inc; originally published by American Society for Biochemistry and Molecular Biology.</rights><rights>2011 by The American Society for Biochemistry and Molecular Biology, Inc. 2011</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c508t-b8aabbc9249c72feb1d04bc2f51777545de79cee16534d9baaa724b498c11f5a3</citedby><cites>FETCH-LOGICAL-c508t-b8aabbc9249c72feb1d04bc2f51777545de79cee16534d9baaa724b498c11f5a3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC3103344/pdf/$$EPDF$$P50$$Gpubmedcentral$$H</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC3103344/$$EHTML$$P50$$Gpubmedcentral$$H</linktohtml><link.rule.ids>230,314,723,776,780,881,27901,27902,53766,53768</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/21474448$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Kumar, Atul</creatorcontrib><creatorcontrib>Saigal, Kashyap</creatorcontrib><creatorcontrib>Malhotra, Ketan</creatorcontrib><creatorcontrib>Sinha, Krishna Murari</creatorcontrib><creatorcontrib>Taneja, Bhupesh</creatorcontrib><title>Structural and Functional Characterization of Rv2966c Protein Reveals an RsmD-like Methyltransferase from Mycobacterium tuberculosis and the Role of Its N-terminal Domain in Target Recognition</title><title>The Journal of biological chemistry</title><addtitle>J Biol Chem</addtitle><description>Nine of ten methylated nucleotides of Escherichia coli 16 S rRNA are conserved in Mycobacterium tuberculosis. All the 10 different methyltransferases are known in E. coli, whereas only TlyA and GidB have been identified in mycobacteria. Here we have identified Rv2966c of M. tuberculosis as an ortholog of RsmD protein of E. coli. We have shown that rv2966c can complement rsmD-deleted E. coli cells. Recombinant Rv2966c can use 30 S ribosomes purified from rsmD-deleted E. coli as substrate and methylate G966 of 16 S rRNA in vitro. Structure determination of the protein shows the protein to be a two-domain structure with a short hairpin domain at the N terminus and a C-terminal domain with the S-adenosylmethionine-MT-fold. We show that the N-terminal hairpin is a minimalist functional domain that helps Rv2966c in target recognition. Deletion of the N-terminal domain prevents binding to nucleic acid substrates, and the truncated protein fails to carry out the m2G966 methylation on 16 S rRNA. The N-terminal domain also binds DNA efficiently, a property that may be utilized under specific conditions of cellular growth.</description><subject>Bacterial Proteins - chemistry</subject><subject>Bacterial Proteins - genetics</subject><subject>Bacterial Proteins - metabolism</subject><subject>Crystal Structure</subject><subject>DNA-binding Protein</subject><subject>Histones - chemistry</subject><subject>Histones - genetics</subject><subject>Histones - metabolism</subject><subject>Methylation</subject><subject>Mycobacterium tuberculosis - enzymology</subject><subject>Mycobacterium tuberculosis - genetics</subject><subject>Protein Structure</subject><subject>Protein Structure and Folding</subject><subject>Protein Structure, Tertiary</subject><subject>Ribosomal RNA (rRNA)</subject><subject>RNA Methylation</subject><subject>RNA Methyltransferase</subject><subject>RNA, Bacterial - chemistry</subject><subject>RNA, Bacterial - genetics</subject><subject>RNA, Bacterial - metabolism</subject><subject>RNA, Ribosomal, 16S - chemistry</subject><subject>RNA, Ribosomal, 16S - genetics</subject><subject>RNA, Ribosomal, 16S - metabolism</subject><subject>RsmD</subject><subject>Rv2966c</subject><subject>S-Adenosylmethionine (SAM)</subject><subject>SAM-MT-fold</subject><subject>Structure-Activity Relationship</subject><subject>tRNA Methyltransferases - chemistry</subject><subject>tRNA Methyltransferases - genetics</subject><subject>tRNA Methyltransferases - metabolism</subject><issn>0021-9258</issn><issn>1083-351X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2011</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp1UUFvFCEYJUZj1-rZm-HmaVpgmJ3hYmK21jbpqllr4o0A880udQZaYDZZf11_moxTGz1ICOSD97334CH0mpITSmp-eqPNyZrmihHCWfMELShpyqKs6PenaEEIo4VgVXOEXsR4Q_Lggj5HR4zymnPeLND91xRGk8ageqxci89HZ5L1LpernQrKJAj2p5qOsO_wZs_Ecmnwl-ATWIc3sAfVx9yKN3E4K3r7A_Aa0u7Qp6Bc7CCoCLgLfsDrg_F6JhwHnEYNwYy9jzb-Vk47wBvfwyRzmSL-VGTkYCcnZ35QWSzPaxW2kLKs8VtnJ1cv0bMuO4BXD_sx-nb-4Xp1UVx9_ni5en9VmIo0qdCNUlobwbgwNetA05ZwbVhX0bquK161UAsDQJdVyVuhlVI145qLxlDaVao8Ru9m3ttRD9AacPmBvbwNdlDhIL2y8t8bZ3dy6_eypKQsOc8Ebx8Igr8bISY52Gig75UDP0bZLAUneWEZeTojTfAxBugeVSiRU-wyxy6n2OUce-5487e5R_yfnDNAzADIX7S3EGQ0FpyB1gYwSbbe_pf8F4A0wjI</recordid><startdate>20110603</startdate><enddate>20110603</enddate><creator>Kumar, Atul</creator><creator>Saigal, Kashyap</creator><creator>Malhotra, Ketan</creator><creator>Sinha, Krishna Murari</creator><creator>Taneja, Bhupesh</creator><general>Elsevier Inc</general><general>American Society for Biochemistry and Molecular Biology</general><scope>6I.</scope><scope>AAFTH</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>20110603</creationdate><title>Structural and Functional Characterization of Rv2966c Protein Reveals an RsmD-like Methyltransferase from Mycobacterium tuberculosis and the Role of Its N-terminal Domain in Target Recognition</title><author>Kumar, Atul ; Saigal, Kashyap ; Malhotra, Ketan ; Sinha, Krishna Murari ; Taneja, Bhupesh</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c508t-b8aabbc9249c72feb1d04bc2f51777545de79cee16534d9baaa724b498c11f5a3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2011</creationdate><topic>Bacterial Proteins - chemistry</topic><topic>Bacterial Proteins - genetics</topic><topic>Bacterial Proteins - metabolism</topic><topic>Crystal Structure</topic><topic>DNA-binding Protein</topic><topic>Histones - chemistry</topic><topic>Histones - genetics</topic><topic>Histones - metabolism</topic><topic>Methylation</topic><topic>Mycobacterium tuberculosis - enzymology</topic><topic>Mycobacterium tuberculosis - genetics</topic><topic>Protein Structure</topic><topic>Protein Structure and Folding</topic><topic>Protein Structure, Tertiary</topic><topic>Ribosomal RNA (rRNA)</topic><topic>RNA Methylation</topic><topic>RNA Methyltransferase</topic><topic>RNA, Bacterial - chemistry</topic><topic>RNA, Bacterial - genetics</topic><topic>RNA, Bacterial - metabolism</topic><topic>RNA, Ribosomal, 16S - chemistry</topic><topic>RNA, Ribosomal, 16S - genetics</topic><topic>RNA, Ribosomal, 16S - metabolism</topic><topic>RsmD</topic><topic>Rv2966c</topic><topic>S-Adenosylmethionine (SAM)</topic><topic>SAM-MT-fold</topic><topic>Structure-Activity Relationship</topic><topic>tRNA Methyltransferases - chemistry</topic><topic>tRNA Methyltransferases - genetics</topic><topic>tRNA Methyltransferases - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Kumar, Atul</creatorcontrib><creatorcontrib>Saigal, Kashyap</creatorcontrib><creatorcontrib>Malhotra, Ketan</creatorcontrib><creatorcontrib>Sinha, Krishna Murari</creatorcontrib><creatorcontrib>Taneja, Bhupesh</creatorcontrib><collection>ScienceDirect Open Access Titles</collection><collection>Elsevier:ScienceDirect:Open Access</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>The Journal of biological chemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Kumar, Atul</au><au>Saigal, Kashyap</au><au>Malhotra, Ketan</au><au>Sinha, Krishna Murari</au><au>Taneja, Bhupesh</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Structural and Functional Characterization of Rv2966c Protein Reveals an RsmD-like Methyltransferase from Mycobacterium tuberculosis and the Role of Its N-terminal Domain in Target Recognition</atitle><jtitle>The Journal of biological chemistry</jtitle><addtitle>J Biol Chem</addtitle><date>2011-06-03</date><risdate>2011</risdate><volume>286</volume><issue>22</issue><spage>19652</spage><epage>19661</epage><pages>19652-19661</pages><issn>0021-9258</issn><eissn>1083-351X</eissn><abstract>Nine of ten methylated nucleotides of Escherichia coli 16 S rRNA are conserved in Mycobacterium tuberculosis. All the 10 different methyltransferases are known in E. coli, whereas only TlyA and GidB have been identified in mycobacteria. Here we have identified Rv2966c of M. tuberculosis as an ortholog of RsmD protein of E. coli. We have shown that rv2966c can complement rsmD-deleted E. coli cells. Recombinant Rv2966c can use 30 S ribosomes purified from rsmD-deleted E. coli as substrate and methylate G966 of 16 S rRNA in vitro. Structure determination of the protein shows the protein to be a two-domain structure with a short hairpin domain at the N terminus and a C-terminal domain with the S-adenosylmethionine-MT-fold. We show that the N-terminal hairpin is a minimalist functional domain that helps Rv2966c in target recognition. Deletion of the N-terminal domain prevents binding to nucleic acid substrates, and the truncated protein fails to carry out the m2G966 methylation on 16 S rRNA. The N-terminal domain also binds DNA efficiently, a property that may be utilized under specific conditions of cellular growth.</abstract><cop>United States</cop><pub>Elsevier Inc</pub><pmid>21474448</pmid><doi>10.1074/jbc.M110.200428</doi><tpages>10</tpages><oa>free_for_read</oa></addata></record> |
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| ispartof | The Journal of biological chemistry, 2011-06, Vol.286 (22), p.19652-19661 |
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| subjects | Bacterial Proteins - chemistry Bacterial Proteins - genetics Bacterial Proteins - metabolism Crystal Structure DNA-binding Protein Histones - chemistry Histones - genetics Histones - metabolism Methylation Mycobacterium tuberculosis - enzymology Mycobacterium tuberculosis - genetics Protein Structure Protein Structure and Folding Protein Structure, Tertiary Ribosomal RNA (rRNA) RNA Methylation RNA Methyltransferase RNA, Bacterial - chemistry RNA, Bacterial - genetics RNA, Bacterial - metabolism RNA, Ribosomal, 16S - chemistry RNA, Ribosomal, 16S - genetics RNA, Ribosomal, 16S - metabolism RsmD Rv2966c S-Adenosylmethionine (SAM) SAM-MT-fold Structure-Activity Relationship tRNA Methyltransferases - chemistry tRNA Methyltransferases - genetics tRNA Methyltransferases - metabolism |
| title | Structural and Functional Characterization of Rv2966c Protein Reveals an RsmD-like Methyltransferase from Mycobacterium tuberculosis and the Role of Its N-terminal Domain in Target Recognition |
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