Isolation, Characterization, and Disruption of the Yeast Gene Encoding Cytosolic NADP-Specific Isocitrate Dehydrogenase
The cytosolic isozyme of NADP-specific isocitrate dehydrogenase (IDP2) was purified from a Saccharomyces cerevisiae mutant containing a chromosomal disruption in the gene encoding the mitochondrial isozyme (IDP1). IDP2 was shown to be a homodimer with a subunit molecular weight of approximately 45 0...
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| Veröffentlicht in: | Biochemistry (Easton) 1994-08, Vol.33 (32), p.9661-9667 |
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| container_issue | 32 |
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| container_title | Biochemistry (Easton) |
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| creator | Loftus, Thomas M Hall, Linda V Anderson, Sondra L McAlister-Henn, Lee |
| description | The cytosolic isozyme of NADP-specific isocitrate dehydrogenase (IDP2) was purified from a Saccharomyces cerevisiae mutant containing a chromosomal disruption in the gene encoding the mitochondrial isozyme (IDP1). IDP2 was shown to be a homodimer with a subunit molecular weight of approximately 45 000 and an isoelectric point of 5.5. Amino acid sequences were obtained for tryptic peptides of IDP2 and used to plan polymerase chain reactions. A resulting 400 bp DNA fragment was used as a hybridization probe to isolate the IDP2 gene from a yeast genomic DNA library. The complete nucleotide sequence of the IDP2 coding region was determined and translated into a 412-residue amino acid sequence. IDP2 and IDP1 were found to be identical in 71% of the aligned residue positions. The identity of the IDP2 gene was confirmed by genomic replacement with a disrupted IDP2 coding region. Haploid yeast strains lacking either or both IDP2 and IDP1 were constructed by genetic crosses of mutant strains containing disruptions in chromosomal IDP2 and IDP1 loci. No dramatic different in growth rates with common carbon sources could be attributed to these disruptions |
| doi_str_mv | 10.1021/bi00198a035 |
| format | Article |
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IDP2 was shown to be a homodimer with a subunit molecular weight of approximately 45 000 and an isoelectric point of 5.5. Amino acid sequences were obtained for tryptic peptides of IDP2 and used to plan polymerase chain reactions. A resulting 400 bp DNA fragment was used as a hybridization probe to isolate the IDP2 gene from a yeast genomic DNA library. The complete nucleotide sequence of the IDP2 coding region was determined and translated into a 412-residue amino acid sequence. IDP2 and IDP1 were found to be identical in 71% of the aligned residue positions. The identity of the IDP2 gene was confirmed by genomic replacement with a disrupted IDP2 coding region. Haploid yeast strains lacking either or both IDP2 and IDP1 were constructed by genetic crosses of mutant strains containing disruptions in chromosomal IDP2 and IDP1 loci. No dramatic different in growth rates with common carbon sources could be attributed to these disruptions</description><identifier>ISSN: 0006-2960</identifier><identifier>EISSN: 1520-4995</identifier><identifier>DOI: 10.1021/bi00198a035</identifier><identifier>PMID: 8068643</identifier><language>eng</language><publisher>United States: American Chemical Society</publisher><subject>Amino Acid Sequence ; Base Sequence ; Cell Compartmentation ; CLONACION ; CLONAGE ; Cloning, Molecular ; COENZIMAS ; COENZYME ; COMPOSICION QUIMICA ; COMPOSITION CHIMIQUE ; Cytosol - enzymology ; GENE ; GENES ; Genes, Fungal - genetics ; Isocitrate Dehydrogenase - genetics ; Isocitrate Dehydrogenase - isolation & purification ; ISOCITRATE DESHYDROGENASE ; ISOCITRATO DESHIDROGENASA ; Isoenzymes - genetics ; Isoenzymes - isolation & purification ; Molecular Sequence Data ; Mutagenesis, Insertional ; MUTANT ; MUTANTES ; NUCLEOTIDE ; NUCLEOTIDOS ; Restriction Mapping ; SACCHAROMYCES CEREVISIAE ; Saccharomyces cerevisiae - enzymology ; Saccharomyces cerevisiae - genetics ; SECUENCIA NUCLEICA ; Sequence Analysis, DNA ; Sequence Homology, Amino Acid ; SEQUENCE NUCLEIQUE</subject><ispartof>Biochemistry (Easton), 1994-08, Vol.33 (32), p.9661-9667</ispartof><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-a470t-e61f18eafe2f43c2ba516ee4e450837df6f800c7a90245589f90c968bc05d303</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://pubs.acs.org/doi/pdf/10.1021/bi00198a035$$EPDF$$P50$$Gacs$$H</linktopdf><linktohtml>$$Uhttps://pubs.acs.org/doi/10.1021/bi00198a035$$EHTML$$P50$$Gacs$$H</linktohtml><link.rule.ids>314,780,784,2765,27076,27924,27925,56738,56788</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/8068643$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Loftus, Thomas M</creatorcontrib><creatorcontrib>Hall, Linda V</creatorcontrib><creatorcontrib>Anderson, Sondra L</creatorcontrib><creatorcontrib>McAlister-Henn, Lee</creatorcontrib><title>Isolation, Characterization, and Disruption of the Yeast Gene Encoding Cytosolic NADP-Specific Isocitrate Dehydrogenase</title><title>Biochemistry (Easton)</title><addtitle>Biochemistry</addtitle><description>The cytosolic isozyme of NADP-specific isocitrate dehydrogenase (IDP2) was purified from a Saccharomyces cerevisiae mutant containing a chromosomal disruption in the gene encoding the mitochondrial isozyme (IDP1). IDP2 was shown to be a homodimer with a subunit molecular weight of approximately 45 000 and an isoelectric point of 5.5. Amino acid sequences were obtained for tryptic peptides of IDP2 and used to plan polymerase chain reactions. A resulting 400 bp DNA fragment was used as a hybridization probe to isolate the IDP2 gene from a yeast genomic DNA library. The complete nucleotide sequence of the IDP2 coding region was determined and translated into a 412-residue amino acid sequence. IDP2 and IDP1 were found to be identical in 71% of the aligned residue positions. The identity of the IDP2 gene was confirmed by genomic replacement with a disrupted IDP2 coding region. Haploid yeast strains lacking either or both IDP2 and IDP1 were constructed by genetic crosses of mutant strains containing disruptions in chromosomal IDP2 and IDP1 loci. No dramatic different in growth rates with common carbon sources could be attributed to these disruptions</description><subject>Amino Acid Sequence</subject><subject>Base Sequence</subject><subject>Cell Compartmentation</subject><subject>CLONACION</subject><subject>CLONAGE</subject><subject>Cloning, Molecular</subject><subject>COENZIMAS</subject><subject>COENZYME</subject><subject>COMPOSICION QUIMICA</subject><subject>COMPOSITION CHIMIQUE</subject><subject>Cytosol - enzymology</subject><subject>GENE</subject><subject>GENES</subject><subject>Genes, Fungal - genetics</subject><subject>Isocitrate Dehydrogenase - genetics</subject><subject>Isocitrate Dehydrogenase - isolation & purification</subject><subject>ISOCITRATE DESHYDROGENASE</subject><subject>ISOCITRATO DESHIDROGENASA</subject><subject>Isoenzymes - genetics</subject><subject>Isoenzymes - isolation & purification</subject><subject>Molecular Sequence Data</subject><subject>Mutagenesis, Insertional</subject><subject>MUTANT</subject><subject>MUTANTES</subject><subject>NUCLEOTIDE</subject><subject>NUCLEOTIDOS</subject><subject>Restriction Mapping</subject><subject>SACCHAROMYCES CEREVISIAE</subject><subject>Saccharomyces cerevisiae - enzymology</subject><subject>Saccharomyces cerevisiae - genetics</subject><subject>SECUENCIA NUCLEICA</subject><subject>Sequence Analysis, DNA</subject><subject>Sequence Homology, Amino Acid</subject><subject>SEQUENCE NUCLEIQUE</subject><issn>0006-2960</issn><issn>1520-4995</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1994</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNptkEFv0zAYhi0EGmVw4oaE5BMcIPA5sZ34OLVjmzTBUMskuFiu87n1aONiO4Ly60lJNXHgZL1-Hz369BLynME7BiV7v_QATDUGKvGATJgooeBKiYdkAgCyKJWEx-RJSndD5FDzE3LSgGwkrybk51UKG5N96N7S6dpEYzNG__v4Y7qWznyK_e6QaXA0r5F-RZMyvcAO6XlnQ-u7FZ3ucxhM3tKPZ7ObYr5D692QBr31OZqMdIbrfRvDCjuT8Cl55Mwm4bPje0oWH84X08vi-tPF1fTsujC8hlygZI41aByWjle2XBrBJCJHLqCp6tZJ1wDY2igouRCNcgqsks3SgmgrqE7Jq1G7i-FHjynrrU8WNxvTYeiTZlLWiv8F34ygjSGliE7vot-auNcM9GFl_c_KA_3yqO2XW2zv2eOsQ1-MvU8Zf93XJn7Xsq5qoRc3cw234vZbdflZ84F_MfLOBG1W0Sf9Za4EV0IcZK_H0tik70Ifu2Gw_571B5qjm-I</recordid><startdate>19940816</startdate><enddate>19940816</enddate><creator>Loftus, Thomas M</creator><creator>Hall, Linda V</creator><creator>Anderson, Sondra L</creator><creator>McAlister-Henn, Lee</creator><general>American Chemical Society</general><scope>FBQ</scope><scope>BSCLL</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>8FD</scope><scope>FR3</scope><scope>M7N</scope><scope>P64</scope><scope>RC3</scope></search><sort><creationdate>19940816</creationdate><title>Isolation, Characterization, and Disruption of the Yeast Gene Encoding Cytosolic NADP-Specific Isocitrate Dehydrogenase</title><author>Loftus, Thomas M ; Hall, Linda V ; Anderson, Sondra L ; McAlister-Henn, Lee</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-a470t-e61f18eafe2f43c2ba516ee4e450837df6f800c7a90245589f90c968bc05d303</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1994</creationdate><topic>Amino Acid Sequence</topic><topic>Base Sequence</topic><topic>Cell Compartmentation</topic><topic>CLONACION</topic><topic>CLONAGE</topic><topic>Cloning, Molecular</topic><topic>COENZIMAS</topic><topic>COENZYME</topic><topic>COMPOSICION QUIMICA</topic><topic>COMPOSITION CHIMIQUE</topic><topic>Cytosol - enzymology</topic><topic>GENE</topic><topic>GENES</topic><topic>Genes, Fungal - genetics</topic><topic>Isocitrate Dehydrogenase - genetics</topic><topic>Isocitrate Dehydrogenase - isolation & purification</topic><topic>ISOCITRATE DESHYDROGENASE</topic><topic>ISOCITRATO DESHIDROGENASA</topic><topic>Isoenzymes - genetics</topic><topic>Isoenzymes - isolation & purification</topic><topic>Molecular Sequence Data</topic><topic>Mutagenesis, Insertional</topic><topic>MUTANT</topic><topic>MUTANTES</topic><topic>NUCLEOTIDE</topic><topic>NUCLEOTIDOS</topic><topic>Restriction Mapping</topic><topic>SACCHAROMYCES CEREVISIAE</topic><topic>Saccharomyces cerevisiae - enzymology</topic><topic>Saccharomyces cerevisiae - genetics</topic><topic>SECUENCIA NUCLEICA</topic><topic>Sequence Analysis, DNA</topic><topic>Sequence Homology, Amino Acid</topic><topic>SEQUENCE NUCLEIQUE</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Loftus, Thomas M</creatorcontrib><creatorcontrib>Hall, Linda V</creatorcontrib><creatorcontrib>Anderson, Sondra L</creatorcontrib><creatorcontrib>McAlister-Henn, Lee</creatorcontrib><collection>AGRIS</collection><collection>Istex</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Technology Research Database</collection><collection>Engineering Research Database</collection><collection>Algology Mycology and Protozoology Abstracts (Microbiology C)</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Genetics Abstracts</collection><jtitle>Biochemistry (Easton)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Loftus, Thomas M</au><au>Hall, Linda V</au><au>Anderson, Sondra L</au><au>McAlister-Henn, Lee</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Isolation, Characterization, and Disruption of the Yeast Gene Encoding Cytosolic NADP-Specific Isocitrate Dehydrogenase</atitle><jtitle>Biochemistry (Easton)</jtitle><addtitle>Biochemistry</addtitle><date>1994-08-16</date><risdate>1994</risdate><volume>33</volume><issue>32</issue><spage>9661</spage><epage>9667</epage><pages>9661-9667</pages><issn>0006-2960</issn><eissn>1520-4995</eissn><abstract>The cytosolic isozyme of NADP-specific isocitrate dehydrogenase (IDP2) was purified from a Saccharomyces cerevisiae mutant containing a chromosomal disruption in the gene encoding the mitochondrial isozyme (IDP1). IDP2 was shown to be a homodimer with a subunit molecular weight of approximately 45 000 and an isoelectric point of 5.5. Amino acid sequences were obtained for tryptic peptides of IDP2 and used to plan polymerase chain reactions. A resulting 400 bp DNA fragment was used as a hybridization probe to isolate the IDP2 gene from a yeast genomic DNA library. The complete nucleotide sequence of the IDP2 coding region was determined and translated into a 412-residue amino acid sequence. IDP2 and IDP1 were found to be identical in 71% of the aligned residue positions. The identity of the IDP2 gene was confirmed by genomic replacement with a disrupted IDP2 coding region. Haploid yeast strains lacking either or both IDP2 and IDP1 were constructed by genetic crosses of mutant strains containing disruptions in chromosomal IDP2 and IDP1 loci. No dramatic different in growth rates with common carbon sources could be attributed to these disruptions</abstract><cop>United States</cop><pub>American Chemical Society</pub><pmid>8068643</pmid><doi>10.1021/bi00198a035</doi><tpages>7</tpages></addata></record> |
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| ispartof | Biochemistry (Easton), 1994-08, Vol.33 (32), p.9661-9667 |
| issn | 0006-2960 1520-4995 |
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| source | MEDLINE; ACS Publications |
| subjects | Amino Acid Sequence Base Sequence Cell Compartmentation CLONACION CLONAGE Cloning, Molecular COENZIMAS COENZYME COMPOSICION QUIMICA COMPOSITION CHIMIQUE Cytosol - enzymology GENE GENES Genes, Fungal - genetics Isocitrate Dehydrogenase - genetics Isocitrate Dehydrogenase - isolation & purification ISOCITRATE DESHYDROGENASE ISOCITRATO DESHIDROGENASA Isoenzymes - genetics Isoenzymes - isolation & purification Molecular Sequence Data Mutagenesis, Insertional MUTANT MUTANTES NUCLEOTIDE NUCLEOTIDOS Restriction Mapping SACCHAROMYCES CEREVISIAE Saccharomyces cerevisiae - enzymology Saccharomyces cerevisiae - genetics SECUENCIA NUCLEICA Sequence Analysis, DNA Sequence Homology, Amino Acid SEQUENCE NUCLEIQUE |
| title | Isolation, Characterization, and Disruption of the Yeast Gene Encoding Cytosolic NADP-Specific Isocitrate Dehydrogenase |
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