Characterization and sequencing of the active site of 1-aminocyclopropane-1-carboxylate synthase
The pyridoxal phosphate (PLP)-dependent 1-aminocyclopropane-1-carboxylic acid (ACC) synthase (S-adenosyl-L-methionine methylthioadenosine-lyase, EC 4.4.1.14), the key enzyme in ethylene biosynthesis, is inactivated by its substrate S-adenosylmethionine (AdoMet). Apple ACC synthase was purified with...
Gespeichert in:
| Veröffentlicht in: | Proceedings of the National Academy of Sciences - PNAS 1990-10, Vol.87 (20), p.7930-7934 |
|---|---|
| Hauptverfasser: | , , , , |
| Format: | Artikel |
| Sprache: | eng |
| Schlagworte: | |
| Online-Zugang: | Volltext |
| Tags: |
Tag hinzufügen
Keine Tags, Fügen Sie den ersten Tag hinzu!
|
| container_end_page | 7934 |
|---|---|
| container_issue | 20 |
| container_start_page | 7930 |
| container_title | Proceedings of the National Academy of Sciences - PNAS |
| container_volume | 87 |
| creator | Yip, W.K. (University of California, Davis, CA) Dong, J.G Kenny, J.W Thompson, G.A Yang, S.F |
| description | The pyridoxal phosphate (PLP)-dependent 1-aminocyclopropane-1-carboxylic acid (ACC) synthase (S-adenosyl-L-methionine methylthioadenosine-lyase, EC 4.4.1.14), the key enzyme in ethylene biosynthesis, is inactivated by its substrate S-adenosylmethionine (AdoMet). Apple ACC synthase was purified with an immunoaffinity gel, and its active site was probed with NaB3H4or Ado[14C]Met. HPLC separation of the trypsin digest yielded a single radioactive peptide. Peptide sequencing of both3H- and14C-labeled peptides revealed a common dodecapeptide of Ser-Leu-Ser-Xaa-Asp-Leu-Gly-Leu-Pro-Gly-Phe-Arg, where Xaa was the modified, radioactive residue in each case. Acid hydrolysis of the3H-labeled enzyme released radioactive N-pyridoxyllysine, indicating that the active-site peptide contained lysine at position 4. Mass spectrometry of the14C-labeled peptide indicated a protonated molecular ion at m/z 1390.6, from which the mass of Xaa was calculated to be 229, a number that is equivalent to the mass of a lysine residue alkylated by the 2-aminobutyrate portion of AdoMet, as we previously proposed. These results indicate that the same active-site lysine binds the PLP and convalently links to the 2-aminobutyrate portion of AdoMet during inactivation. The active site of tomato ACC synthase was probed in the same manner with Ado[14C]Met. Sequencing of the tomato active-site peptide revealed two highly conserved dodecapeptides; the minor peptide possessed a sequence identical to that of the apple enzyme, whereas the major peptide differed from the minor peptide in that methionine replaced leucine at position 6. |
| doi_str_mv | 10.1073/pnas.87.20.7930 |
| format | Article |
| fullrecord | <record><control><sourceid>jstor_proqu</sourceid><recordid>TN_cdi_proquest_miscellaneous_80110589</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><jstor_id>2355448</jstor_id><sourcerecordid>2355448</sourcerecordid><originalsourceid>FETCH-LOGICAL-c538t-d4d1c4431197ad279707a593e27cd79c9066765ede8ed3fddbbd7ae7b15886a43</originalsourceid><addsrcrecordid>eNp9kc-L1DAcxYso67h6FkSlCOqps0mbNAl4kcFfsOBB9xy_TdKZLJ2km2SWHf96U1p33YunQN7nm_e-eUXxHKM1Rqw5Gx3ENWfrGq2ZaNCDYoWRwFVLBHpYrBCqWcVJTR4XT2K8RAgJytFJcVLjuiZErIpfmx0EUMkE-xuS9a4Ep8torg7GKeu2pe_LtDNlRuy1KaNNZrrCFeyt8-qoBj8GP4IzFa4UhM7fHAfIUDy6tINonhaPehiiebacp8XF508_N1-r8-9fvm0-nleKNjxVmmisCGkwFgx0zQRDDKhoTM2UZkIJ1LaspUYbbnTTa911moFhHaact0Ca0-LD_O546PZGK-NSgEGOwe4hHKUHK-8rzu7k1l9LSnhL8_ibedzHZGVUeU-1U945o5KklBPcsgy9WzyCzx8Uk9zbqMww5PX9IUqOMEaUiwyezaAKPsZg-tscGMmpNzn1JjmTNZJTb3ni1b_xb_mlqKy_XXSICoY-QK4n3j0rKGEt55l7vXCTwV_5ntH7_wKyPwxDMjcpky9n8jImH-4SNZQSMjm9mOUevIRtyGkufgiMSMtI8wcw4szI</addsrcrecordid><sourcetype>Open Access Repository</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>80110589</pqid></control><display><type>article</type><title>Characterization and sequencing of the active site of 1-aminocyclopropane-1-carboxylate synthase</title><source>MEDLINE</source><source>Jstor Complete Legacy</source><source>PubMed Central</source><source>Alma/SFX Local Collection</source><source>Free Full-Text Journals in Chemistry</source><creator>Yip, W.K. (University of California, Davis, CA) ; Dong, J.G ; Kenny, J.W ; Thompson, G.A ; Yang, S.F</creator><creatorcontrib>Yip, W.K. (University of California, Davis, CA) ; Dong, J.G ; Kenny, J.W ; Thompson, G.A ; Yang, S.F</creatorcontrib><description>The pyridoxal phosphate (PLP)-dependent 1-aminocyclopropane-1-carboxylic acid (ACC) synthase (S-adenosyl-L-methionine methylthioadenosine-lyase, EC 4.4.1.14), the key enzyme in ethylene biosynthesis, is inactivated by its substrate S-adenosylmethionine (AdoMet). Apple ACC synthase was purified with an immunoaffinity gel, and its active site was probed with NaB3H4or Ado[14C]Met. HPLC separation of the trypsin digest yielded a single radioactive peptide. Peptide sequencing of both3H- and14C-labeled peptides revealed a common dodecapeptide of Ser-Leu-Ser-Xaa-Asp-Leu-Gly-Leu-Pro-Gly-Phe-Arg, where Xaa was the modified, radioactive residue in each case. Acid hydrolysis of the3H-labeled enzyme released radioactive N-pyridoxyllysine, indicating that the active-site peptide contained lysine at position 4. Mass spectrometry of the14C-labeled peptide indicated a protonated molecular ion at m/z 1390.6, from which the mass of Xaa was calculated to be 229, a number that is equivalent to the mass of a lysine residue alkylated by the 2-aminobutyrate portion of AdoMet, as we previously proposed. These results indicate that the same active-site lysine binds the PLP and convalently links to the 2-aminobutyrate portion of AdoMet during inactivation. The active site of tomato ACC synthase was probed in the same manner with Ado[14C]Met. Sequencing of the tomato active-site peptide revealed two highly conserved dodecapeptides; the minor peptide possessed a sequence identical to that of the apple enzyme, whereas the major peptide differed from the minor peptide in that methionine replaced leucine at position 6.</description><identifier>ISSN: 0027-8424</identifier><identifier>EISSN: 1091-6490</identifier><identifier>DOI: 10.1073/pnas.87.20.7930</identifier><identifier>PMID: 2122449</identifier><identifier>CODEN: PNASA6</identifier><language>eng</language><publisher>Washington, DC: National Academy of Sciences of the United States of America</publisher><subject>550201 - Biochemistry- Tracer Techniques ; ACC ; Active sites ; ADENOSYLMETHIONINE ; AGENTES NOCIVOS ; ALKENES ; AMINO ACID SEQUENCE ; AMINO ACIDS ; AMINO COMPOUNDS ; Analytical, structural and metabolic biochemistry ; BASIC BIOLOGICAL SCIENCES ; Binding Sites ; Biological and medical sciences ; BIOSYNTHESIS ; CARBON 14 COMPOUNDS ; CARBOXY-LYASES ; CARBOXYLIC ACIDS ; COENZIMAS ; COENZYME ; COENZYMES ; Complementary DNA ; COMPOSE AMINE ; COMPUESTOS DE AMINA ; DNA SEQUENCING ; ENZYME INHIBITORS ; Enzymes ; Enzymes and enzyme inhibitors ; ETHYLENE ; FACTEUR NUISIBLE ; Fruit ; Fundamental and applied biological sciences. Psychology ; Gels ; GENES ; HYDROCARBONS ; HYDROGEN COMPOUNDS ; INHIBIDORES DE ENZIMAS ; INHIBITEUR D'ENZYME ; INJURIES ; INJURIOUS FACTORS ; LABELLED COMPOUNDS ; LIASAS ; Liquids ; LYASE ; LYASES ; Lyases - genetics ; Lyases - metabolism ; LYCOPERSICON ESCULENTUM ; LYSINE ; Lysine - analogs & derivatives ; Lysine - analysis ; MADURAMIENTO ; MALUS SYLVESTRIS ; MASS SPECTROSCOPY ; Molecular Sequence Data ; MOLECULAR STRUCTURE ; Monoclonal antibodies ; MURISSAGE ; ORGANIC ACIDS ; ORGANIC COMPOUNDS ; Paper chromatography ; Peptide Mapping ; PEPTIDES ; PLANT GROWTH SUBSTANCES ; Plants - enzymology ; PROTEINS ; Pyridoxal - analogs & derivatives ; Pyridoxal - analysis ; PYRIDOXAL PHOSPHATE ; RIPENING ; Sequence Homology, Nucleic Acid ; Sequencing ; SPECTROSCOPY ; STRUCTURAL CHEMICAL ANALYSIS ; SUBSTANCE DE CROISSANCE VEGETALE ; SUSTANCIAS DE CRECIMIENTO VEGETAL ; SYNTHESIS ; TRITIUM COMPOUNDS ; Trypsin</subject><ispartof>Proceedings of the National Academy of Sciences - PNAS, 1990-10, Vol.87 (20), p.7930-7934</ispartof><rights>1991 INIST-CNRS</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c538t-d4d1c4431197ad279707a593e27cd79c9066765ede8ed3fddbbd7ae7b15886a43</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Uhttp://www.pnas.org/content/87/20.cover.gif</thumbnail><linktopdf>$$Uhttps://www.jstor.org/stable/pdf/2355448$$EPDF$$P50$$Gjstor$$H</linktopdf><linktohtml>$$Uhttps://www.jstor.org/stable/2355448$$EHTML$$P50$$Gjstor$$H</linktohtml><link.rule.ids>230,314,723,776,780,799,881,27903,27904,53770,53772,57996,58229</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=19547688$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/2122449$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink><backlink>$$Uhttps://www.osti.gov/biblio/5584167$$D View this record in Osti.gov$$Hfree_for_read</backlink></links><search><creatorcontrib>Yip, W.K. (University of California, Davis, CA)</creatorcontrib><creatorcontrib>Dong, J.G</creatorcontrib><creatorcontrib>Kenny, J.W</creatorcontrib><creatorcontrib>Thompson, G.A</creatorcontrib><creatorcontrib>Yang, S.F</creatorcontrib><title>Characterization and sequencing of the active site of 1-aminocyclopropane-1-carboxylate synthase</title><title>Proceedings of the National Academy of Sciences - PNAS</title><addtitle>Proc Natl Acad Sci U S A</addtitle><description>The pyridoxal phosphate (PLP)-dependent 1-aminocyclopropane-1-carboxylic acid (ACC) synthase (S-adenosyl-L-methionine methylthioadenosine-lyase, EC 4.4.1.14), the key enzyme in ethylene biosynthesis, is inactivated by its substrate S-adenosylmethionine (AdoMet). Apple ACC synthase was purified with an immunoaffinity gel, and its active site was probed with NaB3H4or Ado[14C]Met. HPLC separation of the trypsin digest yielded a single radioactive peptide. Peptide sequencing of both3H- and14C-labeled peptides revealed a common dodecapeptide of Ser-Leu-Ser-Xaa-Asp-Leu-Gly-Leu-Pro-Gly-Phe-Arg, where Xaa was the modified, radioactive residue in each case. Acid hydrolysis of the3H-labeled enzyme released radioactive N-pyridoxyllysine, indicating that the active-site peptide contained lysine at position 4. Mass spectrometry of the14C-labeled peptide indicated a protonated molecular ion at m/z 1390.6, from which the mass of Xaa was calculated to be 229, a number that is equivalent to the mass of a lysine residue alkylated by the 2-aminobutyrate portion of AdoMet, as we previously proposed. These results indicate that the same active-site lysine binds the PLP and convalently links to the 2-aminobutyrate portion of AdoMet during inactivation. The active site of tomato ACC synthase was probed in the same manner with Ado[14C]Met. Sequencing of the tomato active-site peptide revealed two highly conserved dodecapeptides; the minor peptide possessed a sequence identical to that of the apple enzyme, whereas the major peptide differed from the minor peptide in that methionine replaced leucine at position 6.</description><subject>550201 - Biochemistry- Tracer Techniques</subject><subject>ACC</subject><subject>Active sites</subject><subject>ADENOSYLMETHIONINE</subject><subject>AGENTES NOCIVOS</subject><subject>ALKENES</subject><subject>AMINO ACID SEQUENCE</subject><subject>AMINO ACIDS</subject><subject>AMINO COMPOUNDS</subject><subject>Analytical, structural and metabolic biochemistry</subject><subject>BASIC BIOLOGICAL SCIENCES</subject><subject>Binding Sites</subject><subject>Biological and medical sciences</subject><subject>BIOSYNTHESIS</subject><subject>CARBON 14 COMPOUNDS</subject><subject>CARBOXY-LYASES</subject><subject>CARBOXYLIC ACIDS</subject><subject>COENZIMAS</subject><subject>COENZYME</subject><subject>COENZYMES</subject><subject>Complementary DNA</subject><subject>COMPOSE AMINE</subject><subject>COMPUESTOS DE AMINA</subject><subject>DNA SEQUENCING</subject><subject>ENZYME INHIBITORS</subject><subject>Enzymes</subject><subject>Enzymes and enzyme inhibitors</subject><subject>ETHYLENE</subject><subject>FACTEUR NUISIBLE</subject><subject>Fruit</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Gels</subject><subject>GENES</subject><subject>HYDROCARBONS</subject><subject>HYDROGEN COMPOUNDS</subject><subject>INHIBIDORES DE ENZIMAS</subject><subject>INHIBITEUR D'ENZYME</subject><subject>INJURIES</subject><subject>INJURIOUS FACTORS</subject><subject>LABELLED COMPOUNDS</subject><subject>LIASAS</subject><subject>Liquids</subject><subject>LYASE</subject><subject>LYASES</subject><subject>Lyases - genetics</subject><subject>Lyases - metabolism</subject><subject>LYCOPERSICON ESCULENTUM</subject><subject>LYSINE</subject><subject>Lysine - analogs & derivatives</subject><subject>Lysine - analysis</subject><subject>MADURAMIENTO</subject><subject>MALUS SYLVESTRIS</subject><subject>MASS SPECTROSCOPY</subject><subject>Molecular Sequence Data</subject><subject>MOLECULAR STRUCTURE</subject><subject>Monoclonal antibodies</subject><subject>MURISSAGE</subject><subject>ORGANIC ACIDS</subject><subject>ORGANIC COMPOUNDS</subject><subject>Paper chromatography</subject><subject>Peptide Mapping</subject><subject>PEPTIDES</subject><subject>PLANT GROWTH SUBSTANCES</subject><subject>Plants - enzymology</subject><subject>PROTEINS</subject><subject>Pyridoxal - analogs & derivatives</subject><subject>Pyridoxal - analysis</subject><subject>PYRIDOXAL PHOSPHATE</subject><subject>RIPENING</subject><subject>Sequence Homology, Nucleic Acid</subject><subject>Sequencing</subject><subject>SPECTROSCOPY</subject><subject>STRUCTURAL CHEMICAL ANALYSIS</subject><subject>SUBSTANCE DE CROISSANCE VEGETALE</subject><subject>SUSTANCIAS DE CRECIMIENTO VEGETAL</subject><subject>SYNTHESIS</subject><subject>TRITIUM COMPOUNDS</subject><subject>Trypsin</subject><issn>0027-8424</issn><issn>1091-6490</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1990</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp9kc-L1DAcxYso67h6FkSlCOqps0mbNAl4kcFfsOBB9xy_TdKZLJ2km2SWHf96U1p33YunQN7nm_e-eUXxHKM1Rqw5Gx3ENWfrGq2ZaNCDYoWRwFVLBHpYrBCqWcVJTR4XT2K8RAgJytFJcVLjuiZErIpfmx0EUMkE-xuS9a4Ep8torg7GKeu2pe_LtDNlRuy1KaNNZrrCFeyt8-qoBj8GP4IzFa4UhM7fHAfIUDy6tINonhaPehiiebacp8XF508_N1-r8-9fvm0-nleKNjxVmmisCGkwFgx0zQRDDKhoTM2UZkIJ1LaspUYbbnTTa911moFhHaact0Ca0-LD_O546PZGK-NSgEGOwe4hHKUHK-8rzu7k1l9LSnhL8_ibedzHZGVUeU-1U945o5KklBPcsgy9WzyCzx8Uk9zbqMww5PX9IUqOMEaUiwyezaAKPsZg-tscGMmpNzn1JjmTNZJTb3ni1b_xb_mlqKy_XXSICoY-QK4n3j0rKGEt55l7vXCTwV_5ntH7_wKyPwxDMjcpky9n8jImH-4SNZQSMjm9mOUevIRtyGkufgiMSMtI8wcw4szI</recordid><startdate>19901001</startdate><enddate>19901001</enddate><creator>Yip, W.K. (University of California, Davis, CA)</creator><creator>Dong, J.G</creator><creator>Kenny, J.W</creator><creator>Thompson, G.A</creator><creator>Yang, S.F</creator><general>National Academy of Sciences of the United States of America</general><general>National Acad Sciences</general><scope>FBQ</scope><scope>IQODW</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>OTOTI</scope><scope>5PM</scope></search><sort><creationdate>19901001</creationdate><title>Characterization and sequencing of the active site of 1-aminocyclopropane-1-carboxylate synthase</title><author>Yip, W.K. (University of California, Davis, CA) ; Dong, J.G ; Kenny, J.W ; Thompson, G.A ; Yang, S.F</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c538t-d4d1c4431197ad279707a593e27cd79c9066765ede8ed3fddbbd7ae7b15886a43</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1990</creationdate><topic>550201 - Biochemistry- Tracer Techniques</topic><topic>ACC</topic><topic>Active sites</topic><topic>ADENOSYLMETHIONINE</topic><topic>AGENTES NOCIVOS</topic><topic>ALKENES</topic><topic>AMINO ACID SEQUENCE</topic><topic>AMINO ACIDS</topic><topic>AMINO COMPOUNDS</topic><topic>Analytical, structural and metabolic biochemistry</topic><topic>BASIC BIOLOGICAL SCIENCES</topic><topic>Binding Sites</topic><topic>Biological and medical sciences</topic><topic>BIOSYNTHESIS</topic><topic>CARBON 14 COMPOUNDS</topic><topic>CARBOXY-LYASES</topic><topic>CARBOXYLIC ACIDS</topic><topic>COENZIMAS</topic><topic>COENZYME</topic><topic>COENZYMES</topic><topic>Complementary DNA</topic><topic>COMPOSE AMINE</topic><topic>COMPUESTOS DE AMINA</topic><topic>DNA SEQUENCING</topic><topic>ENZYME INHIBITORS</topic><topic>Enzymes</topic><topic>Enzymes and enzyme inhibitors</topic><topic>ETHYLENE</topic><topic>FACTEUR NUISIBLE</topic><topic>Fruit</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Gels</topic><topic>GENES</topic><topic>HYDROCARBONS</topic><topic>HYDROGEN COMPOUNDS</topic><topic>INHIBIDORES DE ENZIMAS</topic><topic>INHIBITEUR D'ENZYME</topic><topic>INJURIES</topic><topic>INJURIOUS FACTORS</topic><topic>LABELLED COMPOUNDS</topic><topic>LIASAS</topic><topic>Liquids</topic><topic>LYASE</topic><topic>LYASES</topic><topic>Lyases - genetics</topic><topic>Lyases - metabolism</topic><topic>LYCOPERSICON ESCULENTUM</topic><topic>LYSINE</topic><topic>Lysine - analogs & derivatives</topic><topic>Lysine - analysis</topic><topic>MADURAMIENTO</topic><topic>MALUS SYLVESTRIS</topic><topic>MASS SPECTROSCOPY</topic><topic>Molecular Sequence Data</topic><topic>MOLECULAR STRUCTURE</topic><topic>Monoclonal antibodies</topic><topic>MURISSAGE</topic><topic>ORGANIC ACIDS</topic><topic>ORGANIC COMPOUNDS</topic><topic>Paper chromatography</topic><topic>Peptide Mapping</topic><topic>PEPTIDES</topic><topic>PLANT GROWTH SUBSTANCES</topic><topic>Plants - enzymology</topic><topic>PROTEINS</topic><topic>Pyridoxal - analogs & derivatives</topic><topic>Pyridoxal - analysis</topic><topic>PYRIDOXAL PHOSPHATE</topic><topic>RIPENING</topic><topic>Sequence Homology, Nucleic Acid</topic><topic>Sequencing</topic><topic>SPECTROSCOPY</topic><topic>STRUCTURAL CHEMICAL ANALYSIS</topic><topic>SUBSTANCE DE CROISSANCE VEGETALE</topic><topic>SUSTANCIAS DE CRECIMIENTO VEGETAL</topic><topic>SYNTHESIS</topic><topic>TRITIUM COMPOUNDS</topic><topic>Trypsin</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Yip, W.K. (University of California, Davis, CA)</creatorcontrib><creatorcontrib>Dong, J.G</creatorcontrib><creatorcontrib>Kenny, J.W</creatorcontrib><creatorcontrib>Thompson, G.A</creatorcontrib><creatorcontrib>Yang, S.F</creatorcontrib><collection>AGRIS</collection><collection>Pascal-Francis</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>OSTI.GOV</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Proceedings of the National Academy of Sciences - PNAS</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Yip, W.K. (University of California, Davis, CA)</au><au>Dong, J.G</au><au>Kenny, J.W</au><au>Thompson, G.A</au><au>Yang, S.F</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Characterization and sequencing of the active site of 1-aminocyclopropane-1-carboxylate synthase</atitle><jtitle>Proceedings of the National Academy of Sciences - PNAS</jtitle><addtitle>Proc Natl Acad Sci U S A</addtitle><date>1990-10-01</date><risdate>1990</risdate><volume>87</volume><issue>20</issue><spage>7930</spage><epage>7934</epage><pages>7930-7934</pages><issn>0027-8424</issn><eissn>1091-6490</eissn><coden>PNASA6</coden><abstract>The pyridoxal phosphate (PLP)-dependent 1-aminocyclopropane-1-carboxylic acid (ACC) synthase (S-adenosyl-L-methionine methylthioadenosine-lyase, EC 4.4.1.14), the key enzyme in ethylene biosynthesis, is inactivated by its substrate S-adenosylmethionine (AdoMet). Apple ACC synthase was purified with an immunoaffinity gel, and its active site was probed with NaB3H4or Ado[14C]Met. HPLC separation of the trypsin digest yielded a single radioactive peptide. Peptide sequencing of both3H- and14C-labeled peptides revealed a common dodecapeptide of Ser-Leu-Ser-Xaa-Asp-Leu-Gly-Leu-Pro-Gly-Phe-Arg, where Xaa was the modified, radioactive residue in each case. Acid hydrolysis of the3H-labeled enzyme released radioactive N-pyridoxyllysine, indicating that the active-site peptide contained lysine at position 4. Mass spectrometry of the14C-labeled peptide indicated a protonated molecular ion at m/z 1390.6, from which the mass of Xaa was calculated to be 229, a number that is equivalent to the mass of a lysine residue alkylated by the 2-aminobutyrate portion of AdoMet, as we previously proposed. These results indicate that the same active-site lysine binds the PLP and convalently links to the 2-aminobutyrate portion of AdoMet during inactivation. The active site of tomato ACC synthase was probed in the same manner with Ado[14C]Met. Sequencing of the tomato active-site peptide revealed two highly conserved dodecapeptides; the minor peptide possessed a sequence identical to that of the apple enzyme, whereas the major peptide differed from the minor peptide in that methionine replaced leucine at position 6.</abstract><cop>Washington, DC</cop><pub>National Academy of Sciences of the United States of America</pub><pmid>2122449</pmid><doi>10.1073/pnas.87.20.7930</doi><tpages>5</tpages><oa>free_for_read</oa></addata></record> |
| fulltext | fulltext |
| identifier | ISSN: 0027-8424 |
| ispartof | Proceedings of the National Academy of Sciences - PNAS, 1990-10, Vol.87 (20), p.7930-7934 |
| issn | 0027-8424 1091-6490 |
| language | eng |
| recordid | cdi_proquest_miscellaneous_80110589 |
| source | MEDLINE; Jstor Complete Legacy; PubMed Central; Alma/SFX Local Collection; Free Full-Text Journals in Chemistry |
| subjects | 550201 - Biochemistry- Tracer Techniques ACC Active sites ADENOSYLMETHIONINE AGENTES NOCIVOS ALKENES AMINO ACID SEQUENCE AMINO ACIDS AMINO COMPOUNDS Analytical, structural and metabolic biochemistry BASIC BIOLOGICAL SCIENCES Binding Sites Biological and medical sciences BIOSYNTHESIS CARBON 14 COMPOUNDS CARBOXY-LYASES CARBOXYLIC ACIDS COENZIMAS COENZYME COENZYMES Complementary DNA COMPOSE AMINE COMPUESTOS DE AMINA DNA SEQUENCING ENZYME INHIBITORS Enzymes Enzymes and enzyme inhibitors ETHYLENE FACTEUR NUISIBLE Fruit Fundamental and applied biological sciences. Psychology Gels GENES HYDROCARBONS HYDROGEN COMPOUNDS INHIBIDORES DE ENZIMAS INHIBITEUR D'ENZYME INJURIES INJURIOUS FACTORS LABELLED COMPOUNDS LIASAS Liquids LYASE LYASES Lyases - genetics Lyases - metabolism LYCOPERSICON ESCULENTUM LYSINE Lysine - analogs & derivatives Lysine - analysis MADURAMIENTO MALUS SYLVESTRIS MASS SPECTROSCOPY Molecular Sequence Data MOLECULAR STRUCTURE Monoclonal antibodies MURISSAGE ORGANIC ACIDS ORGANIC COMPOUNDS Paper chromatography Peptide Mapping PEPTIDES PLANT GROWTH SUBSTANCES Plants - enzymology PROTEINS Pyridoxal - analogs & derivatives Pyridoxal - analysis PYRIDOXAL PHOSPHATE RIPENING Sequence Homology, Nucleic Acid Sequencing SPECTROSCOPY STRUCTURAL CHEMICAL ANALYSIS SUBSTANCE DE CROISSANCE VEGETALE SUSTANCIAS DE CRECIMIENTO VEGETAL SYNTHESIS TRITIUM COMPOUNDS Trypsin |
| title | Characterization and sequencing of the active site of 1-aminocyclopropane-1-carboxylate synthase |
| url | https://sfx.bib-bvb.de/sfx_tum?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2025-01-23T04%3A19%3A23IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-jstor_proqu&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=Characterization%20and%20sequencing%20of%20the%20active%20site%20of%201-aminocyclopropane-1-carboxylate%20synthase&rft.jtitle=Proceedings%20of%20the%20National%20Academy%20of%20Sciences%20-%20PNAS&rft.au=Yip,%20W.K.%20(University%20of%20California,%20Davis,%20CA)&rft.date=1990-10-01&rft.volume=87&rft.issue=20&rft.spage=7930&rft.epage=7934&rft.pages=7930-7934&rft.issn=0027-8424&rft.eissn=1091-6490&rft.coden=PNASA6&rft_id=info:doi/10.1073/pnas.87.20.7930&rft_dat=%3Cjstor_proqu%3E2355448%3C/jstor_proqu%3E%3Curl%3E%3C/url%3E&disable_directlink=true&sfx.directlink=off&sfx.report_link=0&rft_id=info:oai/&rft_pqid=80110589&rft_id=info:pmid/2122449&rft_jstor_id=2355448&rfr_iscdi=true |