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Conversion of Trp 62 of Hen Egg-White Lysozyme to Tyr by Site-Directed Mutagenesis

An expression plasmid for hen egg-white lysozyme in Saccharomyces cercvisiae was constructed by inserting almost full-length cDNA (about 600 base pairs) encoding hen egg-white pre-lysozyme into a yeast expression vector, pAM 82. The hen lysozyme was expressed under the control of the repressible aci...

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Published in:	Journal of biochemistry (Tokyo) 1987-10, Vol.102 (4), p.733-740
Main Authors:	KUMAGAI, Izumi, KOJIMA, Shuichi, TAMAKI, Eisuke, MIURA, Kin-ichiro
Format:	Article
Language:	English
Subjects:	Animals Base Sequence Biological and medical sciences Chickens Classical genetics, quantitative genetics, hybrids Cloning, Molecular Fundamental and applied biological sciences. Psychology Genetics of eukaryotes. Biological and molecular evolution Molecular Sequence Data Muramidase - genetics Mutation Saccharomyces cerevisiae - genetics Thallophyta, bryophyta Tryptophan Tyrosine Vegetals
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cited_by	cdi_FETCH-LOGICAL-c526t-bed8a09fef03add7c080d0429c676594bbc5c438e2df5008fd364b9ec267e1f63
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container_end_page	740
container_issue	4
container_start_page	733
container_title	Journal of biochemistry (Tokyo)
container_volume	102
creator	KUMAGAI, Izumi KOJIMA, Shuichi TAMAKI, Eisuke MIURA, Kin-ichiro
description	An expression plasmid for hen egg-white lysozyme in Saccharomyces cercvisiae was constructed by inserting almost full-length cDNA (about 600 base pairs) encoding hen egg-white pre-lysozyme into a yeast expression vector, pAM 82. The hen lysozyme was expressed under the control of the repressible acid phosphatase promoter of pAM 82 in S. cerevisiae. About half of the expressed lysozyme was secreted in the yeast growth medium as a precise mature protein which exhibited specific activity consistent with that of authentic hen egg-white lysozyme. The replacement of Trp 62 of hen egg-white lysozyme with a tyrosine residue was performed by site-directed mutagenesis using a 19-mer oligodeoxyribonucleotide. The mutant lysozyme with Tyr 62 was found to exhibit enhanced bacteriolytic activity.
doi_str_mv	10.1093/oxfordjournals.jbchem.a122111
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The hen lysozyme was expressed under the control of the repressible acid phosphatase promoter of pAM 82 in S. cerevisiae. About half of the expressed lysozyme was secreted in the yeast growth medium as a precise mature protein which exhibited specific activity consistent with that of authentic hen egg-white lysozyme. The replacement of Trp 62 of hen egg-white lysozyme with a tyrosine residue was performed by site-directed mutagenesis using a 19-mer oligodeoxyribonucleotide. 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The hen lysozyme was expressed under the control of the repressible acid phosphatase promoter of pAM 82 in S. cerevisiae. About half of the expressed lysozyme was secreted in the yeast growth medium as a precise mature protein which exhibited specific activity consistent with that of authentic hen egg-white lysozyme. The replacement of Trp 62 of hen egg-white lysozyme with a tyrosine residue was performed by site-directed mutagenesis using a 19-mer oligodeoxyribonucleotide. The mutant lysozyme with Tyr 62 was found to exhibit enhanced bacteriolytic activity.</description><subject>Animals</subject><subject>Base Sequence</subject><subject>Biological and medical sciences</subject><subject>Chickens</subject><subject>Classical genetics, quantitative genetics, hybrids</subject><subject>Cloning, Molecular</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Genetics of eukaryotes. Biological and molecular evolution</subject><subject>Molecular Sequence Data</subject><subject>Muramidase - genetics</subject><subject>Mutation</subject><subject>Saccharomyces cerevisiae - genetics</subject><subject>Thallophyta, bryophyta</subject><subject>Tryptophan</subject><subject>Tyrosine</subject><subject>Vegetals</subject><issn>0021-924X</issn><issn>1756-2651</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1987</creationdate><recordtype>article</recordtype><recordid>eNpVkEtLAzEUhYMoWh8_QchCl1PzmGRmFi60VStUBK0obkImualT7aQkU3H89U5pKbi6j3PO5fIhdE5Jn5KCX_gf54Od-WWo9Vfsz0rzAfO-poxRSndQj2ZCJkwKuot6hDCaFCx9O0CHMc5WI-N8H-1zzoQgtIeeBr7-hhArX2Pv8CQssGSrbgQ1vplOk9ePqgE8bqP_beeAG48nbcBli5-7fTKsApgGLH5YNnoKNcQqHqM9130GJ5t6hF5ubyaDUTJ-vLsfXI0TI5hskhJsrknhwBGurc0MyYklKSuMzKQo0rI0wqQ8B2adICR3lsu0LMAwmQF1kh-hy_VdE3yMAZxahGquQ6soUStU6j8qtUalNqi6_Ok6v1iWc7Db9IZNp59tdB2N_nJB16aKW1smU5nTorMla1sVG_jZyjp8KpnxTKjR27u6JU_XZMiHasL_ABJCiLA</recordid><startdate>19871001</startdate><enddate>19871001</enddate><creator>KUMAGAI, Izumi</creator><creator>KOJIMA, Shuichi</creator><creator>TAMAKI, Eisuke</creator><creator>MIURA, Kin-ichiro</creator><general>Oxford University Press</general><scope>BSCLL</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></search><sort><creationdate>19871001</creationdate><title>Conversion of Trp 62 of Hen Egg-White Lysozyme to Tyr by Site-Directed Mutagenesis</title><author>KUMAGAI, Izumi ; KOJIMA, Shuichi ; TAMAKI, Eisuke ; MIURA, Kin-ichiro</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c526t-bed8a09fef03add7c080d0429c676594bbc5c438e2df5008fd364b9ec267e1f63</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1987</creationdate><topic>Animals</topic><topic>Base Sequence</topic><topic>Biological and medical sciences</topic><topic>Chickens</topic><topic>Classical genetics, quantitative genetics, hybrids</topic><topic>Cloning, Molecular</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Genetics of eukaryotes. Biological and molecular evolution</topic><topic>Molecular Sequence Data</topic><topic>Muramidase - genetics</topic><topic>Mutation</topic><topic>Saccharomyces cerevisiae - genetics</topic><topic>Thallophyta, bryophyta</topic><topic>Tryptophan</topic><topic>Tyrosine</topic><topic>Vegetals</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>KUMAGAI, Izumi</creatorcontrib><creatorcontrib>KOJIMA, Shuichi</creatorcontrib><creatorcontrib>TAMAKI, Eisuke</creatorcontrib><creatorcontrib>MIURA, Kin-ichiro</creatorcontrib><collection>Istex</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><jtitle>Journal of biochemistry (Tokyo)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>KUMAGAI, Izumi</au><au>KOJIMA, Shuichi</au><au>TAMAKI, Eisuke</au><au>MIURA, Kin-ichiro</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Conversion of Trp 62 of Hen Egg-White Lysozyme to Tyr by Site-Directed Mutagenesis</atitle><jtitle>Journal of biochemistry (Tokyo)</jtitle><addtitle>J Biochem</addtitle><date>1987-10-01</date><risdate>1987</risdate><volume>102</volume><issue>4</issue><spage>733</spage><epage>740</epage><pages>733-740</pages><issn>0021-924X</issn><eissn>1756-2651</eissn><coden>JOBIAO</coden><abstract>An expression plasmid for hen egg-white lysozyme in Saccharomyces cercvisiae was constructed by inserting almost full-length cDNA (about 600 base pairs) encoding hen egg-white pre-lysozyme into a yeast expression vector, pAM 82. The hen lysozyme was expressed under the control of the repressible acid phosphatase promoter of pAM 82 in S. cerevisiae. About half of the expressed lysozyme was secreted in the yeast growth medium as a precise mature protein which exhibited specific activity consistent with that of authentic hen egg-white lysozyme. The replacement of Trp 62 of hen egg-white lysozyme with a tyrosine residue was performed by site-directed mutagenesis using a 19-mer oligodeoxyribonucleotide. The mutant lysozyme with Tyr 62 was found to exhibit enhanced bacteriolytic activity.</abstract><cop>Oxford</cop><pub>Oxford University Press</pub><pmid>3325501</pmid><doi>10.1093/oxfordjournals.jbchem.a122111</doi><tpages>8</tpages></addata></record>
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source	J-STAGE Free; Oxford University Press:Jisc Collections:Oxford Journal Archive: Access period 2024-2025
subjects	Animals Base Sequence Biological and medical sciences Chickens Classical genetics, quantitative genetics, hybrids Cloning, Molecular Fundamental and applied biological sciences. Psychology Genetics of eukaryotes. Biological and molecular evolution Molecular Sequence Data Muramidase - genetics Mutation Saccharomyces cerevisiae - genetics Thallophyta, bryophyta Tryptophan Tyrosine Vegetals
title	Conversion of Trp 62 of Hen Egg-White Lysozyme to Tyr by Site-Directed Mutagenesis
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