Molecular Cloning, Purification and Functional Implications of Recombinant GST Tagged hGMCSF Cytokine
We report the cloning, purification and cell proliferative activity of a novel recombinant GST tagged human granulocyte macrophage colony stimulating factor (GST-hGMCSF). The hGMCSF gene was PCR amplified from the cDNA of ACHN renal carcinoma cells and was cloned into the bacterial expression vector...
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| Veröffentlicht in: | Applied biochemistry and biotechnology 2013-03, Vol.169 (5), p.1713-1726 |
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| creator | Chaubey, Nidhi Ghosh, Siddhartha Sankar |
| description | We report the cloning, purification and cell proliferative activity of a novel recombinant GST tagged human granulocyte macrophage colony stimulating factor (GST-hGMCSF). The hGMCSF gene was PCR amplified from the cDNA of ACHN renal carcinoma cells and was cloned into the bacterial expression vector. The GST-hGMCSF was purified to homogeneity using glutathione agarose affinity chromatography and subsequently characterized by Western blot, circular dichroism (CD) and MALDI TOF-TOF analysis. Homology modelling studies revealed the possible binding domains of the recombinant cytokine with cognate receptor. The proliferation of THP-1, Raw 264.7, MCF-7 and U87MG cells upon GST-hGMCSF addition was found to be dose dependent. Hence, this functionally active recombinant cytokine has potential application in cancer therapy for stimulating facile growth recovery of normal cell population. |
| doi_str_mv | 10.1007/s12010-012-0052-7 |
| format | Article |
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The hGMCSF gene was PCR amplified from the cDNA of ACHN renal carcinoma cells and was cloned into the bacterial expression vector. The GST-hGMCSF was purified to homogeneity using glutathione agarose affinity chromatography and subsequently characterized by Western blot, circular dichroism (CD) and MALDI TOF-TOF analysis. Homology modelling studies revealed the possible binding domains of the recombinant cytokine with cognate receptor. The proliferation of THP-1, Raw 264.7, MCF-7 and U87MG cells upon GST-hGMCSF addition was found to be dose dependent. Hence, this functionally active recombinant cytokine has potential application in cancer therapy for stimulating facile growth recovery of normal cell population.</description><identifier>ISSN: 0273-2289</identifier><identifier>EISSN: 1559-0291</identifier><identifier>DOI: 10.1007/s12010-012-0052-7</identifier><identifier>PMID: 23334834</identifier><identifier>CODEN: ABIBDL</identifier><language>eng</language><publisher>New York: Springer-Verlag</publisher><subject>Amino Acid Sequence ; Biochemistry ; Biological activity ; Biological and medical sciences ; Biotechnology ; Cancer therapies ; Cell growth ; Cell Line, Tumor ; Cell Proliferation - drug effects ; Cell Survival - drug effects ; Chemistry ; Chemistry and Materials Science ; Cloning ; Cloning, Molecular ; Cytokines ; DNA, Complementary - genetics ; DNA, Complementary - metabolism ; Dose-Response Relationship, Drug ; E coli ; Escherichia coli - genetics ; Fundamental and applied biological sciences. Psychology ; Gene expression ; Granulocyte-Macrophage Colony-Stimulating Factor - chemistry ; Granulocyte-Macrophage Colony-Stimulating Factor - genetics ; Granulocyte-Macrophage Colony-Stimulating Factor - pharmacology ; Granulocytes ; Humans ; Molecular Docking Simulation ; Molecular Dynamics Simulation ; Molecular Sequence Data ; Protein Structure, Secondary ; Protein Structure, Tertiary ; Proteins ; Receptors, Granulocyte-Macrophage Colony-Stimulating Factor - chemistry ; Recombinant Fusion Proteins - chemistry ; Recombinant Fusion Proteins - genetics ; Recombinant Fusion Proteins - pharmacology ; Structural Homology, Protein</subject><ispartof>Applied biochemistry and biotechnology, 2013-03, Vol.169 (5), p.1713-1726</ispartof><rights>Springer Science+Business Media New York 2013</rights><rights>2014 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c435t-45bcb2a6d73682a0b2aad4375bd652403fba3b42056988705d222c80b6aa432d3</citedby><cites>FETCH-LOGICAL-c435t-45bcb2a6d73682a0b2aad4375bd652403fba3b42056988705d222c80b6aa432d3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://link.springer.com/content/pdf/10.1007/s12010-012-0052-7$$EPDF$$P50$$Gspringer$$H</linktopdf><linktohtml>$$Uhttps://link.springer.com/10.1007/s12010-012-0052-7$$EHTML$$P50$$Gspringer$$H</linktohtml><link.rule.ids>314,776,780,27903,27904,41467,42536,51297</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=27566846$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/23334834$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Chaubey, Nidhi</creatorcontrib><creatorcontrib>Ghosh, Siddhartha Sankar</creatorcontrib><title>Molecular Cloning, Purification and Functional Implications of Recombinant GST Tagged hGMCSF Cytokine</title><title>Applied biochemistry and biotechnology</title><addtitle>Appl Biochem Biotechnol</addtitle><addtitle>Appl Biochem Biotechnol</addtitle><description>We report the cloning, purification and cell proliferative activity of a novel recombinant GST tagged human granulocyte macrophage colony stimulating factor (GST-hGMCSF). The hGMCSF gene was PCR amplified from the cDNA of ACHN renal carcinoma cells and was cloned into the bacterial expression vector. The GST-hGMCSF was purified to homogeneity using glutathione agarose affinity chromatography and subsequently characterized by Western blot, circular dichroism (CD) and MALDI TOF-TOF analysis. Homology modelling studies revealed the possible binding domains of the recombinant cytokine with cognate receptor. The proliferation of THP-1, Raw 264.7, MCF-7 and U87MG cells upon GST-hGMCSF addition was found to be dose dependent. Hence, this functionally active recombinant cytokine has potential application in cancer therapy for stimulating facile growth recovery of normal cell population.</description><subject>Amino Acid Sequence</subject><subject>Biochemistry</subject><subject>Biological activity</subject><subject>Biological and medical sciences</subject><subject>Biotechnology</subject><subject>Cancer therapies</subject><subject>Cell growth</subject><subject>Cell Line, Tumor</subject><subject>Cell Proliferation - drug effects</subject><subject>Cell Survival - drug effects</subject><subject>Chemistry</subject><subject>Chemistry and Materials Science</subject><subject>Cloning</subject><subject>Cloning, Molecular</subject><subject>Cytokines</subject><subject>DNA, Complementary - genetics</subject><subject>DNA, Complementary - metabolism</subject><subject>Dose-Response Relationship, Drug</subject><subject>E coli</subject><subject>Escherichia coli - genetics</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Gene expression</subject><subject>Granulocyte-Macrophage Colony-Stimulating Factor - chemistry</subject><subject>Granulocyte-Macrophage Colony-Stimulating Factor - genetics</subject><subject>Granulocyte-Macrophage Colony-Stimulating Factor - pharmacology</subject><subject>Granulocytes</subject><subject>Humans</subject><subject>Molecular Docking Simulation</subject><subject>Molecular Dynamics Simulation</subject><subject>Molecular Sequence Data</subject><subject>Protein Structure, Secondary</subject><subject>Protein Structure, Tertiary</subject><subject>Proteins</subject><subject>Receptors, Granulocyte-Macrophage Colony-Stimulating Factor - chemistry</subject><subject>Recombinant Fusion Proteins - chemistry</subject><subject>Recombinant Fusion Proteins - genetics</subject><subject>Recombinant Fusion Proteins - pharmacology</subject><subject>Structural Homology, Protein</subject><issn>0273-2289</issn><issn>1559-0291</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2013</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><recordid>eNqNkV2L1DAUhoMo7jj6A7yRgAheWD05aT7mUoozLuyiuON1SNN07Noms0l7sf_elhk_EASvknCevOckDyHPGbxlAOpdZggMCmBYAAgs1AOyYkJsCsANe0hWgIoXiHpzQZ7kfAszqIV6TC6Qc15qXq6Iv469d1NvE636GLpweEM_T6lrO2fHLgZqQ0O3U3DLwfb0cjj251KmsaVfvItD3QUbRrq72dO9PRx8Q7_trqubLa3ux_i9C_4pedTaPvtn53VNvm4_7KuPxdWn3WX1_qpwJRdjUYra1Whlo7jUaGHe26bkStSNFFgCb2vL6xJByI3WCkSDiE5DLa0tOTZ8TV6fco8p3k0-j2bosvN9b4OPUzaMI9eAIOV_oEyC0FyoGX35F3obpzT_xkIJxVDJebY1YSfKpZhz8q05pm6w6d4wMIsuc9JlZgtm0WWW5Bfn5KkefPPrxk8_M_DqDNjsbN8mG1yXf3NKSKnL5TV44vJcCgef_hjxn91_ABidqgM</recordid><startdate>20130301</startdate><enddate>20130301</enddate><creator>Chaubey, Nidhi</creator><creator>Ghosh, Siddhartha Sankar</creator><general>Springer-Verlag</general><general>Springer</general><general>Springer Nature B.V</general><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>3V.</scope><scope>7ST</scope><scope>7T7</scope><scope>7TM</scope><scope>7X7</scope><scope>7XB</scope><scope>88A</scope><scope>88E</scope><scope>88I</scope><scope>8AO</scope><scope>8FD</scope><scope>8FE</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>ABUWG</scope><scope>AEUYN</scope><scope>AFKRA</scope><scope>AZQEC</scope><scope>BBNVY</scope><scope>BENPR</scope><scope>BHPHI</scope><scope>C1K</scope><scope>CCPQU</scope><scope>DWQXO</scope><scope>FR3</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>LK8</scope><scope>M0S</scope><scope>M1P</scope><scope>M2P</scope><scope>M7P</scope><scope>P64</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>Q9U</scope><scope>RC3</scope><scope>SOI</scope><scope>7X8</scope><scope>7QO</scope><scope>7T5</scope><scope>H94</scope></search><sort><creationdate>20130301</creationdate><title>Molecular Cloning, Purification and Functional Implications of Recombinant GST Tagged hGMCSF Cytokine</title><author>Chaubey, Nidhi ; Ghosh, Siddhartha Sankar</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c435t-45bcb2a6d73682a0b2aad4375bd652403fba3b42056988705d222c80b6aa432d3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2013</creationdate><topic>Amino Acid Sequence</topic><topic>Biochemistry</topic><topic>Biological activity</topic><topic>Biological and medical sciences</topic><topic>Biotechnology</topic><topic>Cancer therapies</topic><topic>Cell growth</topic><topic>Cell Line, Tumor</topic><topic>Cell Proliferation - drug effects</topic><topic>Cell Survival - drug effects</topic><topic>Chemistry</topic><topic>Chemistry and Materials Science</topic><topic>Cloning</topic><topic>Cloning, Molecular</topic><topic>Cytokines</topic><topic>DNA, Complementary - genetics</topic><topic>DNA, Complementary - metabolism</topic><topic>Dose-Response Relationship, Drug</topic><topic>E coli</topic><topic>Escherichia coli - genetics</topic><topic>Fundamental and applied biological sciences. 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The hGMCSF gene was PCR amplified from the cDNA of ACHN renal carcinoma cells and was cloned into the bacterial expression vector. The GST-hGMCSF was purified to homogeneity using glutathione agarose affinity chromatography and subsequently characterized by Western blot, circular dichroism (CD) and MALDI TOF-TOF analysis. Homology modelling studies revealed the possible binding domains of the recombinant cytokine with cognate receptor. The proliferation of THP-1, Raw 264.7, MCF-7 and U87MG cells upon GST-hGMCSF addition was found to be dose dependent. Hence, this functionally active recombinant cytokine has potential application in cancer therapy for stimulating facile growth recovery of normal cell population.</abstract><cop>New York</cop><pub>Springer-Verlag</pub><pmid>23334834</pmid><doi>10.1007/s12010-012-0052-7</doi><tpages>14</tpages></addata></record> |
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| identifier | ISSN: 0273-2289 |
| ispartof | Applied biochemistry and biotechnology, 2013-03, Vol.169 (5), p.1713-1726 |
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| subjects | Amino Acid Sequence Biochemistry Biological activity Biological and medical sciences Biotechnology Cancer therapies Cell growth Cell Line, Tumor Cell Proliferation - drug effects Cell Survival - drug effects Chemistry Chemistry and Materials Science Cloning Cloning, Molecular Cytokines DNA, Complementary - genetics DNA, Complementary - metabolism Dose-Response Relationship, Drug E coli Escherichia coli - genetics Fundamental and applied biological sciences. Psychology Gene expression Granulocyte-Macrophage Colony-Stimulating Factor - chemistry Granulocyte-Macrophage Colony-Stimulating Factor - genetics Granulocyte-Macrophage Colony-Stimulating Factor - pharmacology Granulocytes Humans Molecular Docking Simulation Molecular Dynamics Simulation Molecular Sequence Data Protein Structure, Secondary Protein Structure, Tertiary Proteins Receptors, Granulocyte-Macrophage Colony-Stimulating Factor - chemistry Recombinant Fusion Proteins - chemistry Recombinant Fusion Proteins - genetics Recombinant Fusion Proteins - pharmacology Structural Homology, Protein |
| title | Molecular Cloning, Purification and Functional Implications of Recombinant GST Tagged hGMCSF Cytokine |
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