Dbp9p, a Member of the DEAD Box Protein Family, Exhibits DNA Helicase Activity
The yeast Dbp9p is a member of the DEAD box family of RNA helicases, which are thought to be involved in RNA metabolism. Dbp9p seems to function in ribosomal RNA biogenesis, but it has not been biochemically characterized. To analyze the enzymatic characteristics of the protein, we expressed a recom...
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| Veröffentlicht in: | The Journal of biological chemistry 2004-05, Vol.279 (20), p.20692-20698 |
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| container_issue | 20 |
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| container_title | The Journal of biological chemistry |
| container_volume | 279 |
| creator | Kikuma, Takashi Ohtsu, Masaya Utsugi, Takahiko Koga, Shoko Okuhara, Kohji Eki, Toshihiko Fujimori, Fumihiro Murakami, Yasufumi |
| description | The yeast Dbp9p is a member of the DEAD box family of RNA helicases, which are thought to be involved in RNA metabolism. Dbp9p
seems to function in ribosomal RNA biogenesis, but it has not been biochemically characterized. To analyze the enzymatic characteristics
of the protein, we expressed a recombinant Dbp9p in Escherichia coli and purified it to homogeneity. The purified protein exhibited RNA unwinding and binding activity in the absence of NTP,
and this activity was abolished by a mutation in the RNA-binding domain. We then characterized the ATPase activity of Dbp9p
with respect to cofactor specificity; the activity was found to be severely inhibited by yeast total RNA and moderately inhibited
by poly(U), poly(A), and poly(C) but to be stimulated by yeast genomic DNA and salmon sperm DNA. In addition, Dbp9p exhibited
DNA-DNA and DNA-RNA helicase activity in the presence of ATP. These results indicate that Dbp9p has biochemical characteristics
unique among DEAD box proteins. |
| doi_str_mv | 10.1074/jbc.M400231200 |
| format | Article |
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seems to function in ribosomal RNA biogenesis, but it has not been biochemically characterized. To analyze the enzymatic characteristics
of the protein, we expressed a recombinant Dbp9p in Escherichia coli and purified it to homogeneity. The purified protein exhibited RNA unwinding and binding activity in the absence of NTP,
and this activity was abolished by a mutation in the RNA-binding domain. We then characterized the ATPase activity of Dbp9p
with respect to cofactor specificity; the activity was found to be severely inhibited by yeast total RNA and moderately inhibited
by poly(U), poly(A), and poly(C) but to be stimulated by yeast genomic DNA and salmon sperm DNA. In addition, Dbp9p exhibited
DNA-DNA and DNA-RNA helicase activity in the presence of ATP. These results indicate that Dbp9p has biochemical characteristics
unique among DEAD box proteins.</description><identifier>ISSN: 0021-9258</identifier><identifier>EISSN: 1083-351X</identifier><identifier>DOI: 10.1074/jbc.M400231200</identifier><identifier>PMID: 15028736</identifier><language>eng</language><publisher>United States: American Society for Biochemistry and Molecular Biology</publisher><subject>Adenosine Triphosphatases - metabolism ; Base Sequence ; DEAD-box RNA Helicases ; DNA Helicases - isolation & purification ; DNA Helicases - metabolism ; DNA, Fungal - chemistry ; DNA, Fungal - genetics ; DNA, Fungal - metabolism ; DNA, Single-Stranded - drug effects ; DNA, Single-Stranded - metabolism ; Kinetics ; Nuclear Proteins - isolation & purification ; Nuclear Proteins - metabolism ; RNA Helicases - isolation & purification ; RNA Helicases - metabolism ; RNA, Fungal - metabolism ; Saccharomyces cerevisiae ; Saccharomyces cerevisiae - enzymology ; Saccharomyces cerevisiae - genetics ; Saccharomyces cerevisiae Proteins - isolation & purification ; Saccharomyces cerevisiae Proteins - metabolism</subject><ispartof>The Journal of biological chemistry, 2004-05, Vol.279 (20), p.20692-20698</ispartof><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c457t-20d898b3ca36e5ba6933ec581b4f5ffedab37b9d95a020c1de513153ace655e43</citedby><cites>FETCH-LOGICAL-c457t-20d898b3ca36e5ba6933ec581b4f5ffedab37b9d95a020c1de513153ace655e43</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,27924,27925</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/15028736$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Kikuma, Takashi</creatorcontrib><creatorcontrib>Ohtsu, Masaya</creatorcontrib><creatorcontrib>Utsugi, Takahiko</creatorcontrib><creatorcontrib>Koga, Shoko</creatorcontrib><creatorcontrib>Okuhara, Kohji</creatorcontrib><creatorcontrib>Eki, Toshihiko</creatorcontrib><creatorcontrib>Fujimori, Fumihiro</creatorcontrib><creatorcontrib>Murakami, Yasufumi</creatorcontrib><title>Dbp9p, a Member of the DEAD Box Protein Family, Exhibits DNA Helicase Activity</title><title>The Journal of biological chemistry</title><addtitle>J Biol Chem</addtitle><description>The yeast Dbp9p is a member of the DEAD box family of RNA helicases, which are thought to be involved in RNA metabolism. Dbp9p
seems to function in ribosomal RNA biogenesis, but it has not been biochemically characterized. To analyze the enzymatic characteristics
of the protein, we expressed a recombinant Dbp9p in Escherichia coli and purified it to homogeneity. The purified protein exhibited RNA unwinding and binding activity in the absence of NTP,
and this activity was abolished by a mutation in the RNA-binding domain. We then characterized the ATPase activity of Dbp9p
with respect to cofactor specificity; the activity was found to be severely inhibited by yeast total RNA and moderately inhibited
by poly(U), poly(A), and poly(C) but to be stimulated by yeast genomic DNA and salmon sperm DNA. In addition, Dbp9p exhibited
DNA-DNA and DNA-RNA helicase activity in the presence of ATP. These results indicate that Dbp9p has biochemical characteristics
unique among DEAD box proteins.</description><subject>Adenosine Triphosphatases - metabolism</subject><subject>Base Sequence</subject><subject>DEAD-box RNA Helicases</subject><subject>DNA Helicases - isolation & purification</subject><subject>DNA Helicases - metabolism</subject><subject>DNA, Fungal - chemistry</subject><subject>DNA, Fungal - genetics</subject><subject>DNA, Fungal - metabolism</subject><subject>DNA, Single-Stranded - drug effects</subject><subject>DNA, Single-Stranded - metabolism</subject><subject>Kinetics</subject><subject>Nuclear Proteins - isolation & purification</subject><subject>Nuclear Proteins - metabolism</subject><subject>RNA Helicases - isolation & purification</subject><subject>RNA Helicases - metabolism</subject><subject>RNA, Fungal - metabolism</subject><subject>Saccharomyces cerevisiae</subject><subject>Saccharomyces cerevisiae - enzymology</subject><subject>Saccharomyces cerevisiae - genetics</subject><subject>Saccharomyces cerevisiae Proteins - isolation & purification</subject><subject>Saccharomyces cerevisiae Proteins - metabolism</subject><issn>0021-9258</issn><issn>1083-351X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2004</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpFkE1Lw0AQhhdRbK1ePcoexFNT9yObZI-1H1ZoqwcFb8vuZmK2JE3Nptr-eyMtdBgYGJ73PTwI3VIyoCQOH1fGDhYhIYxTRsgZ6lKS8IAL-nmOuu2bBpKJpIOuvF-RdkJJL1GHCsKSmEddtBybjdz0scYLKA3UuMpwkwMeT4Zj_FTt8FtdNeDWeKpLV-z7eLLLnXGNx-PlEM-gcFZ7wEPbuB_X7K_RRaYLDzfH20Mf08n7aBbMX59fRsN5YEMRNwEjaSITw63mEQijI8k5WJFQE2YiyyDVhsdGplJowoilKQjKqeDaQiQEhLyHHg69m7r63oJvVOm8haLQa6i2XtFYxm0yasHBAbR15X0NmdrUrtT1XlGi_g2q1qA6GWwDd8fmrSkhPeFHZS1wfwBy95X_uhqUcZXNoVQsloqRdiPJ-B8-x3Wr</recordid><startdate>20040514</startdate><enddate>20040514</enddate><creator>Kikuma, Takashi</creator><creator>Ohtsu, Masaya</creator><creator>Utsugi, Takahiko</creator><creator>Koga, Shoko</creator><creator>Okuhara, Kohji</creator><creator>Eki, Toshihiko</creator><creator>Fujimori, Fumihiro</creator><creator>Murakami, Yasufumi</creator><general>American Society for Biochemistry and Molecular Biology</general><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>7TM</scope><scope>M7N</scope></search><sort><creationdate>20040514</creationdate><title>Dbp9p, a Member of the DEAD Box Protein Family, Exhibits DNA Helicase Activity</title><author>Kikuma, Takashi ; Ohtsu, Masaya ; Utsugi, Takahiko ; Koga, Shoko ; Okuhara, Kohji ; Eki, Toshihiko ; Fujimori, Fumihiro ; Murakami, Yasufumi</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c457t-20d898b3ca36e5ba6933ec581b4f5ffedab37b9d95a020c1de513153ace655e43</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2004</creationdate><topic>Adenosine Triphosphatases - metabolism</topic><topic>Base Sequence</topic><topic>DEAD-box RNA Helicases</topic><topic>DNA Helicases - isolation & purification</topic><topic>DNA Helicases - metabolism</topic><topic>DNA, Fungal - chemistry</topic><topic>DNA, Fungal - genetics</topic><topic>DNA, Fungal - metabolism</topic><topic>DNA, Single-Stranded - drug effects</topic><topic>DNA, Single-Stranded - metabolism</topic><topic>Kinetics</topic><topic>Nuclear Proteins - isolation & purification</topic><topic>Nuclear Proteins - metabolism</topic><topic>RNA Helicases - isolation & purification</topic><topic>RNA Helicases - metabolism</topic><topic>RNA, Fungal - metabolism</topic><topic>Saccharomyces cerevisiae</topic><topic>Saccharomyces cerevisiae - enzymology</topic><topic>Saccharomyces cerevisiae - genetics</topic><topic>Saccharomyces cerevisiae Proteins - isolation & purification</topic><topic>Saccharomyces cerevisiae Proteins - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Kikuma, Takashi</creatorcontrib><creatorcontrib>Ohtsu, Masaya</creatorcontrib><creatorcontrib>Utsugi, Takahiko</creatorcontrib><creatorcontrib>Koga, Shoko</creatorcontrib><creatorcontrib>Okuhara, Kohji</creatorcontrib><creatorcontrib>Eki, Toshihiko</creatorcontrib><creatorcontrib>Fujimori, Fumihiro</creatorcontrib><creatorcontrib>Murakami, Yasufumi</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Nucleic Acids Abstracts</collection><collection>Algology Mycology and Protozoology Abstracts (Microbiology C)</collection><jtitle>The Journal of biological chemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Kikuma, Takashi</au><au>Ohtsu, Masaya</au><au>Utsugi, Takahiko</au><au>Koga, Shoko</au><au>Okuhara, Kohji</au><au>Eki, Toshihiko</au><au>Fujimori, Fumihiro</au><au>Murakami, Yasufumi</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Dbp9p, a Member of the DEAD Box Protein Family, Exhibits DNA Helicase Activity</atitle><jtitle>The Journal of biological chemistry</jtitle><addtitle>J Biol Chem</addtitle><date>2004-05-14</date><risdate>2004</risdate><volume>279</volume><issue>20</issue><spage>20692</spage><epage>20698</epage><pages>20692-20698</pages><issn>0021-9258</issn><eissn>1083-351X</eissn><abstract>The yeast Dbp9p is a member of the DEAD box family of RNA helicases, which are thought to be involved in RNA metabolism. Dbp9p
seems to function in ribosomal RNA biogenesis, but it has not been biochemically characterized. To analyze the enzymatic characteristics
of the protein, we expressed a recombinant Dbp9p in Escherichia coli and purified it to homogeneity. The purified protein exhibited RNA unwinding and binding activity in the absence of NTP,
and this activity was abolished by a mutation in the RNA-binding domain. We then characterized the ATPase activity of Dbp9p
with respect to cofactor specificity; the activity was found to be severely inhibited by yeast total RNA and moderately inhibited
by poly(U), poly(A), and poly(C) but to be stimulated by yeast genomic DNA and salmon sperm DNA. In addition, Dbp9p exhibited
DNA-DNA and DNA-RNA helicase activity in the presence of ATP. These results indicate that Dbp9p has biochemical characteristics
unique among DEAD box proteins.</abstract><cop>United States</cop><pub>American Society for Biochemistry and Molecular Biology</pub><pmid>15028736</pmid><doi>10.1074/jbc.M400231200</doi><tpages>7</tpages><oa>free_for_read</oa></addata></record> |
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| ispartof | The Journal of biological chemistry, 2004-05, Vol.279 (20), p.20692-20698 |
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| language | eng |
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| source | MEDLINE; EZB-FREE-00999 freely available EZB journals; Alma/SFX Local Collection |
| subjects | Adenosine Triphosphatases - metabolism Base Sequence DEAD-box RNA Helicases DNA Helicases - isolation & purification DNA Helicases - metabolism DNA, Fungal - chemistry DNA, Fungal - genetics DNA, Fungal - metabolism DNA, Single-Stranded - drug effects DNA, Single-Stranded - metabolism Kinetics Nuclear Proteins - isolation & purification Nuclear Proteins - metabolism RNA Helicases - isolation & purification RNA Helicases - metabolism RNA, Fungal - metabolism Saccharomyces cerevisiae Saccharomyces cerevisiae - enzymology Saccharomyces cerevisiae - genetics Saccharomyces cerevisiae Proteins - isolation & purification Saccharomyces cerevisiae Proteins - metabolism |
| title | Dbp9p, a Member of the DEAD Box Protein Family, Exhibits DNA Helicase Activity |
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