Biochemical characterization of an ectonucleotide pyrophosphatase/phosphodiesterase (E-NPP, E.C. 3.1.4.1) from rat cardiac soluble and microsomal fractions
In this study, we have reported the kinetic and biochemical characterization of ectonucleotide pyrophosphatase/phosphodiesterase (E-NPP) activity in rat cardiac fractions, one soluble and the other enriched in vesicles derived from sarcoplasmic reticulum. Both fractions demonstrated E-NPP activities...
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| Veröffentlicht in: | Journal of enzyme inhibition and medicinal chemistry 2012-02, Vol.27 (1), p.29-36 |
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| creator | Pochmann, Daniela Innocente, Adrine Maria Buffon, Andréia Freitas Sarkis, João José Porciúncula, Lisiane De Oliveira |
| description | In this study, we have reported the kinetic and biochemical characterization of ectonucleotide pyrophosphatase/phosphodiesterase (E-NPP) activity in rat cardiac fractions, one soluble and the other enriched in vesicles derived from sarcoplasmic reticulum. Both fractions demonstrated E-NPP activities, which could be observed by extracellular hydrolysis of p-nitrophenyl-5′-thymidine monophosphate (p-Nph-5′-TMP) and other biochemical characteristics. The KM values for the hydrolysis of p-Nph-5′-TMP in soluble and microsomal fractions were 118.53 ± 27.28 and 91.92 ± 12.49 µM, respectively. The Vmax values calculated were 2.56 ± 0.15 and 113.87 ± 21.09 nmol p-nitrophenol/min/mg of protein in soluble and microsomal fractions, respectively. Among the compounds tested to evaluate the possible activity of other enzymes on p-Nph-5′-TMP hydrolysis, only suramin (0.25 mM) produced a significant inhibition of substrate hydrolysis. Thus, our results strongly suggest the presence of E-NPP enzymes in subcellular fractions of rat heart, which could be involved in nucleotide signalling in the cardiac tissue. |
| doi_str_mv | 10.3109/14756366.2011.574129 |
| format | Article |
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Both fractions demonstrated E-NPP activities, which could be observed by extracellular hydrolysis of p-nitrophenyl-5′-thymidine monophosphate (p-Nph-5′-TMP) and other biochemical characteristics. The KM values for the hydrolysis of p-Nph-5′-TMP in soluble and microsomal fractions were 118.53 ± 27.28 and 91.92 ± 12.49 µM, respectively. The Vmax values calculated were 2.56 ± 0.15 and 113.87 ± 21.09 nmol p-nitrophenol/min/mg of protein in soluble and microsomal fractions, respectively. Among the compounds tested to evaluate the possible activity of other enzymes on p-Nph-5′-TMP hydrolysis, only suramin (0.25 mM) produced a significant inhibition of substrate hydrolysis. Thus, our results strongly suggest the presence of E-NPP enzymes in subcellular fractions of rat heart, which could be involved in nucleotide signalling in the cardiac tissue.</description><identifier>ISSN: 1475-6366</identifier><identifier>EISSN: 1475-6374</identifier><identifier>DOI: 10.3109/14756366.2011.574129</identifier><identifier>PMID: 21534863</identifier><language>eng</language><publisher>England: Informa Healthcare</publisher><subject>Adenine nucleotides ; Animals ; E-NPPs ; Heart Ventricles - enzymology ; Hydrolysis ; Male ; microsomes ; Microsomes - enzymology ; Phosphoric Diester Hydrolases - metabolism ; Pyrophosphatases - antagonists & inhibitors ; Pyrophosphatases - metabolism ; rat heart ; Rats ; Rats, Wistar ; Sarcoplasmic Reticulum - enzymology ; Solubility ; Structure-Activity Relationship ; Suramin - pharmacology</subject><ispartof>Journal of enzyme inhibition and medicinal chemistry, 2012-02, Vol.27 (1), p.29-36</ispartof><rights>2012 Informa UK, Ltd. 2012</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c417t-97a1b33d37120e0e29faac2e6e786fc823a0e468bdb8c4429cb047ea4e6aa1273</citedby><cites>FETCH-LOGICAL-c417t-97a1b33d37120e0e29faac2e6e786fc823a0e468bdb8c4429cb047ea4e6aa1273</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,776,780,27901,27902</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/21534863$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Pochmann, Daniela</creatorcontrib><creatorcontrib>Innocente, Adrine Maria</creatorcontrib><creatorcontrib>Buffon, Andréia</creatorcontrib><creatorcontrib>Freitas Sarkis, João José</creatorcontrib><creatorcontrib>Porciúncula, Lisiane De Oliveira</creatorcontrib><title>Biochemical characterization of an ectonucleotide pyrophosphatase/phosphodiesterase (E-NPP, E.C. 3.1.4.1) from rat cardiac soluble and microsomal fractions</title><title>Journal of enzyme inhibition and medicinal chemistry</title><addtitle>J Enzyme Inhib Med Chem</addtitle><description>In this study, we have reported the kinetic and biochemical characterization of ectonucleotide pyrophosphatase/phosphodiesterase (E-NPP) activity in rat cardiac fractions, one soluble and the other enriched in vesicles derived from sarcoplasmic reticulum. Both fractions demonstrated E-NPP activities, which could be observed by extracellular hydrolysis of p-nitrophenyl-5′-thymidine monophosphate (p-Nph-5′-TMP) and other biochemical characteristics. The KM values for the hydrolysis of p-Nph-5′-TMP in soluble and microsomal fractions were 118.53 ± 27.28 and 91.92 ± 12.49 µM, respectively. The Vmax values calculated were 2.56 ± 0.15 and 113.87 ± 21.09 nmol p-nitrophenol/min/mg of protein in soluble and microsomal fractions, respectively. Among the compounds tested to evaluate the possible activity of other enzymes on p-Nph-5′-TMP hydrolysis, only suramin (0.25 mM) produced a significant inhibition of substrate hydrolysis. Thus, our results strongly suggest the presence of E-NPP enzymes in subcellular fractions of rat heart, which could be involved in nucleotide signalling in the cardiac tissue.</description><subject>Adenine nucleotides</subject><subject>Animals</subject><subject>E-NPPs</subject><subject>Heart Ventricles - enzymology</subject><subject>Hydrolysis</subject><subject>Male</subject><subject>microsomes</subject><subject>Microsomes - enzymology</subject><subject>Phosphoric Diester Hydrolases - metabolism</subject><subject>Pyrophosphatases - antagonists & inhibitors</subject><subject>Pyrophosphatases - metabolism</subject><subject>rat heart</subject><subject>Rats</subject><subject>Rats, Wistar</subject><subject>Sarcoplasmic Reticulum - enzymology</subject><subject>Solubility</subject><subject>Structure-Activity Relationship</subject><subject>Suramin - pharmacology</subject><issn>1475-6366</issn><issn>1475-6374</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2012</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp9Uctu1TAUjBCIlsIfIOQdIJHUr-skmyJ6dXlIFXQBa-vEOVFcOXGwE6HbX-FncZS2EpuufHQ0c2Y8k2WvGS0Eo_U5k-VOCaUKThkrdqVkvH6Sna7rXIlSPn2YlTrJXsR4QylnnMnn2QlnOyErJU6zv5fWmx4Ha8AR00MAM2OwtzBbPxLfERgJmtmPi3HoZ9simY7BT72PUw8zRDzfZt9ajImaNuTdIf9-ff2BHIp9QUTBClmw96QLfiABZmIgtBYMid4tjcMk0ZJkIPjoh-SiWz0k9fgye9aBi_jq7j3Lfn0-_Nx_za9-fPm2_3SVG8nKOa9LYI0QrSgZp0iR1x2A4aiwrFRnKi6AolRV0zaVkZLXpqGyRJCoABgvxVn2drs7Bf97Sb_Qg40GnYMR_RJ1zRKpLmmVkHJDrmZjwE5PwQ4QjppRvbai71vRayt6ayXR3twJLM2A7QPpvoYE-LgB7Nj5MMAfH1yrZzg6H1Ico7FxPf-oxMV_F3oEN_cpadQ3fgljyu9xj_8A-jGw6w</recordid><startdate>201202</startdate><enddate>201202</enddate><creator>Pochmann, Daniela</creator><creator>Innocente, Adrine Maria</creator><creator>Buffon, Andréia</creator><creator>Freitas Sarkis, João José</creator><creator>Porciúncula, Lisiane De Oliveira</creator><general>Informa Healthcare</general><general>Taylor & Francis</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>7X8</scope></search><sort><creationdate>201202</creationdate><title>Biochemical characterization of an ectonucleotide pyrophosphatase/phosphodiesterase (E-NPP, E.C. 3.1.4.1) from rat cardiac soluble and microsomal fractions</title><author>Pochmann, Daniela ; Innocente, Adrine Maria ; Buffon, Andréia ; Freitas Sarkis, João José ; Porciúncula, Lisiane De Oliveira</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c417t-97a1b33d37120e0e29faac2e6e786fc823a0e468bdb8c4429cb047ea4e6aa1273</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2012</creationdate><topic>Adenine nucleotides</topic><topic>Animals</topic><topic>E-NPPs</topic><topic>Heart Ventricles - enzymology</topic><topic>Hydrolysis</topic><topic>Male</topic><topic>microsomes</topic><topic>Microsomes - enzymology</topic><topic>Phosphoric Diester Hydrolases - metabolism</topic><topic>Pyrophosphatases - antagonists & inhibitors</topic><topic>Pyrophosphatases - metabolism</topic><topic>rat heart</topic><topic>Rats</topic><topic>Rats, Wistar</topic><topic>Sarcoplasmic Reticulum - enzymology</topic><topic>Solubility</topic><topic>Structure-Activity Relationship</topic><topic>Suramin - pharmacology</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Pochmann, Daniela</creatorcontrib><creatorcontrib>Innocente, Adrine Maria</creatorcontrib><creatorcontrib>Buffon, Andréia</creatorcontrib><creatorcontrib>Freitas Sarkis, João José</creatorcontrib><creatorcontrib>Porciúncula, Lisiane De Oliveira</creatorcontrib><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><jtitle>Journal of enzyme inhibition and medicinal chemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Pochmann, Daniela</au><au>Innocente, Adrine Maria</au><au>Buffon, Andréia</au><au>Freitas Sarkis, João José</au><au>Porciúncula, Lisiane De Oliveira</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Biochemical characterization of an ectonucleotide pyrophosphatase/phosphodiesterase (E-NPP, E.C. 3.1.4.1) from rat cardiac soluble and microsomal fractions</atitle><jtitle>Journal of enzyme inhibition and medicinal chemistry</jtitle><addtitle>J Enzyme Inhib Med Chem</addtitle><date>2012-02</date><risdate>2012</risdate><volume>27</volume><issue>1</issue><spage>29</spage><epage>36</epage><pages>29-36</pages><issn>1475-6366</issn><eissn>1475-6374</eissn><abstract>In this study, we have reported the kinetic and biochemical characterization of ectonucleotide pyrophosphatase/phosphodiesterase (E-NPP) activity in rat cardiac fractions, one soluble and the other enriched in vesicles derived from sarcoplasmic reticulum. Both fractions demonstrated E-NPP activities, which could be observed by extracellular hydrolysis of p-nitrophenyl-5′-thymidine monophosphate (p-Nph-5′-TMP) and other biochemical characteristics. The KM values for the hydrolysis of p-Nph-5′-TMP in soluble and microsomal fractions were 118.53 ± 27.28 and 91.92 ± 12.49 µM, respectively. The Vmax values calculated were 2.56 ± 0.15 and 113.87 ± 21.09 nmol p-nitrophenol/min/mg of protein in soluble and microsomal fractions, respectively. Among the compounds tested to evaluate the possible activity of other enzymes on p-Nph-5′-TMP hydrolysis, only suramin (0.25 mM) produced a significant inhibition of substrate hydrolysis. Thus, our results strongly suggest the presence of E-NPP enzymes in subcellular fractions of rat heart, which could be involved in nucleotide signalling in the cardiac tissue.</abstract><cop>England</cop><pub>Informa Healthcare</pub><pmid>21534863</pmid><doi>10.3109/14756366.2011.574129</doi><tpages>8</tpages></addata></record> |
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| subjects | Adenine nucleotides Animals E-NPPs Heart Ventricles - enzymology Hydrolysis Male microsomes Microsomes - enzymology Phosphoric Diester Hydrolases - metabolism Pyrophosphatases - antagonists & inhibitors Pyrophosphatases - metabolism rat heart Rats Rats, Wistar Sarcoplasmic Reticulum - enzymology Solubility Structure-Activity Relationship Suramin - pharmacology |
| title | Biochemical characterization of an ectonucleotide pyrophosphatase/phosphodiesterase (E-NPP, E.C. 3.1.4.1) from rat cardiac soluble and microsomal fractions |
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