Autophosphorylation of the Insulin-like Growth Factor I Receptor Cytoplasmic Domain
The cytoplasmic domain of the β subunit of the insulin-like growth factor I receptor (amino acids 936–1337) was overexpressed in Sf9 insect cells using a baculovirus expression system, and the 6-His tagged receptor was purified by metal-affinity chromatography. Autophosphorylation of the receptor wa...
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| Veröffentlicht in: | Biochemical and biophysical research communications 2000-12, Vol.279 (3), p.955-960 |
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| container_title | Biochemical and biophysical research communications |
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| creator | Lopaczynski, Wlodzimierz Terry, Cheryl Nissley, Peter |
| description | The cytoplasmic domain of the β subunit of the insulin-like growth factor I receptor (amino acids 936–1337) was overexpressed in Sf9 insect cells using a baculovirus expression system, and the 6-His tagged receptor was purified by metal-affinity chromatography. Autophosphorylation of the receptor was concentration dependent, consistent with a trans phosphorylation mechanism. Phosphoamino acid analysis of the autophosphorylated receptor showed predominantly phosphotyrosine, but phosphoserine and phosphothreonine were also present. However, when the receptor was further purified by gel filtration on Sephadex G-100 and then autophosphorylated, phosphoaminoacid analysis showed only phosphotyrosine. We conclude that the IGF-I receptor tyrosine kinase is not a dual-specificity kinase and that autophosphorylation of the β subunit is by a trans mechanism. |
| doi_str_mv | 10.1006/bbrc.2000.4046 |
| format | Article |
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Autophosphorylation of the receptor was concentration dependent, consistent with a trans phosphorylation mechanism. Phosphoamino acid analysis of the autophosphorylated receptor showed predominantly phosphotyrosine, but phosphoserine and phosphothreonine were also present. However, when the receptor was further purified by gel filtration on Sephadex G-100 and then autophosphorylated, phosphoaminoacid analysis showed only phosphotyrosine. 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Autophosphorylation of the receptor was concentration dependent, consistent with a trans phosphorylation mechanism. Phosphoamino acid analysis of the autophosphorylated receptor showed predominantly phosphotyrosine, but phosphoserine and phosphothreonine were also present. However, when the receptor was further purified by gel filtration on Sephadex G-100 and then autophosphorylated, phosphoaminoacid analysis showed only phosphotyrosine. We conclude that the IGF-I receptor tyrosine kinase is not a dual-specificity kinase and that autophosphorylation of the β subunit is by a trans mechanism.</description><subject>6-His tag</subject><subject>Animals</subject><subject>Cells, Cultured</subject><subject>Chromatography, Gel</subject><subject>Cytoplasm - metabolism</subject><subject>Dimerization</subject><subject>Disulfides - metabolism</subject><subject>Humans</subject><subject>IGF-I receptor dimer</subject><subject>Insecta</subject><subject>insulin receptor</subject><subject>Phosphorylation</subject><subject>Plasma - metabolism</subject><subject>Protein Structure, Tertiary</subject><subject>Protein-Serine-Threonine Kinases - metabolism</subject><subject>Rabbits</subject><subject>Receptor, IGF Type 1 - chemistry</subject><subject>Receptor, IGF Type 1 - metabolism</subject><subject>Serine - metabolism</subject><subject>Threonine - metabolism</subject><issn>0006-291X</issn><issn>1090-2104</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2000</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp1kE1Lw0AQhhdRbK1ePUpO3lJnk-xucizV1kJB8AO8LZvNhK4m2bibKP33JrTgycMww_DMC_MQck1hTgH4XZ47PY8AYJ5Awk_IlEIGYUQhOSXTYc3DKKPvE3Lh_QcApQnPzsmEUsqjhPEpeVn0nW131g_l9pXqjG0CWwbdDoNN4_vKNGFlPjFYO_vT7YKV0p11wSZ4Ro3tOC73Q0ClfG10cG9rZZpLclaqyuPVsc_I2-rhdfkYbp_Wm-ViG-o4gS5M87QUmWA8j4s4zbTSKgclGPJCMEgxB10IrUWEMRNplGNZ0DRjiJhqlmAUz8jtIbd19qtH38naeI1VpRq0vZciYpwKxgZwfgC1s947LGXrTK3cXlKQo0Y5apSjRjlqHA5ujsl9XmPxhx-9DUB6AHD479ugk14bbDQWxqHuZGHNf9m_WgmCdA</recordid><startdate>20001229</startdate><enddate>20001229</enddate><creator>Lopaczynski, Wlodzimierz</creator><creator>Terry, Cheryl</creator><creator>Nissley, Peter</creator><general>Elsevier Inc</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>20001229</creationdate><title>Autophosphorylation of the Insulin-like Growth Factor I Receptor Cytoplasmic Domain</title><author>Lopaczynski, Wlodzimierz ; Terry, Cheryl ; Nissley, Peter</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c340t-8b8f79756b3d389cacab0a75e6d7508eb0cd7cc72e35782befd1895eee8c54e23</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2000</creationdate><topic>6-His tag</topic><topic>Animals</topic><topic>Cells, Cultured</topic><topic>Chromatography, Gel</topic><topic>Cytoplasm - metabolism</topic><topic>Dimerization</topic><topic>Disulfides - metabolism</topic><topic>Humans</topic><topic>IGF-I receptor dimer</topic><topic>Insecta</topic><topic>insulin receptor</topic><topic>Phosphorylation</topic><topic>Plasma - metabolism</topic><topic>Protein Structure, Tertiary</topic><topic>Protein-Serine-Threonine Kinases - metabolism</topic><topic>Rabbits</topic><topic>Receptor, IGF Type 1 - chemistry</topic><topic>Receptor, IGF Type 1 - metabolism</topic><topic>Serine - metabolism</topic><topic>Threonine - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Lopaczynski, Wlodzimierz</creatorcontrib><creatorcontrib>Terry, Cheryl</creatorcontrib><creatorcontrib>Nissley, Peter</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>Biochemical and biophysical research communications</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Lopaczynski, Wlodzimierz</au><au>Terry, Cheryl</au><au>Nissley, Peter</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Autophosphorylation of the Insulin-like Growth Factor I Receptor Cytoplasmic Domain</atitle><jtitle>Biochemical and biophysical research communications</jtitle><addtitle>Biochem Biophys Res Commun</addtitle><date>2000-12-29</date><risdate>2000</risdate><volume>279</volume><issue>3</issue><spage>955</spage><epage>960</epage><pages>955-960</pages><issn>0006-291X</issn><eissn>1090-2104</eissn><abstract>The cytoplasmic domain of the β subunit of the insulin-like growth factor I receptor (amino acids 936–1337) was overexpressed in Sf9 insect cells using a baculovirus expression system, and the 6-His tagged receptor was purified by metal-affinity chromatography. Autophosphorylation of the receptor was concentration dependent, consistent with a trans phosphorylation mechanism. Phosphoamino acid analysis of the autophosphorylated receptor showed predominantly phosphotyrosine, but phosphoserine and phosphothreonine were also present. However, when the receptor was further purified by gel filtration on Sephadex G-100 and then autophosphorylated, phosphoaminoacid analysis showed only phosphotyrosine. We conclude that the IGF-I receptor tyrosine kinase is not a dual-specificity kinase and that autophosphorylation of the β subunit is by a trans mechanism.</abstract><cop>United States</cop><pub>Elsevier Inc</pub><pmid>11162456</pmid><doi>10.1006/bbrc.2000.4046</doi><tpages>6</tpages></addata></record> |
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| ispartof | Biochemical and biophysical research communications, 2000-12, Vol.279 (3), p.955-960 |
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| subjects | 6-His tag Animals Cells, Cultured Chromatography, Gel Cytoplasm - metabolism Dimerization Disulfides - metabolism Humans IGF-I receptor dimer Insecta insulin receptor Phosphorylation Plasma - metabolism Protein Structure, Tertiary Protein-Serine-Threonine Kinases - metabolism Rabbits Receptor, IGF Type 1 - chemistry Receptor, IGF Type 1 - metabolism Serine - metabolism Threonine - metabolism |
| title | Autophosphorylation of the Insulin-like Growth Factor I Receptor Cytoplasmic Domain |
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