In silico and experimental improvement of bacteriorhodopsin production in Halobacterium salinarum R1 by increasing DNA-binding affinity of Bat through Q661R/Q665R substitutions in HTH motif
DNA-binding motif of bacterioopsin activator (Bat) protein is a Helix–Turn–Helix motif, which binds to bop promoter and induces bacterioopsin (Bop) expression under light and low oxygen tension. Bacterioopsin is linked to retinal to produce bacteriorhodopsin (BR), which in turn supplies energy sourc...
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| Veröffentlicht in: | Extremophiles : life under extreme conditions 2019-01, Vol.23 (1), p.59-67 |
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| creator | Mirfeizollahi, Azadeh Yakhchali, Bagher Deldar, Ali Asghar Karkhane, Ali Asghar |
| description | DNA-binding motif of bacterioopsin activator (Bat) protein is a Helix–Turn–Helix motif, which binds to
bop
promoter and induces bacterioopsin (Bop) expression under light and low oxygen tension. Bacterioopsin is linked to retinal to produce bacteriorhodopsin (BR), which in turn supplies energy source in
Halobacterium salinarum
. In this study, effect of Bat HTH motif–promoter DNA interaction on bacterioopsin (Bop) expression was investigated using in silico and experimental approaches. Molecular docking showed that the most stable DNA–protein complex was generated by Q661R/Q665R mutant. Based on the in silico analysis, HTH motif was mutated using site-directed mutagenesis and
Hbt. salinarum
recombinant strains were developed by introduction of mutant
bat
genes. Double positively charged amino acid substitutions (Q661R/Q665R) in second helix of HTH motif increased whereas deletion of this region decreased BR production. However, other single substitutions (Q665R and Q661H) did not change BR production. These findings represent key role of HTH motif stability for DNA binding and regulation of bacterioopsin (Bop) expression and bacteriorhodopsin (BR) production independent of environmental condition. |
| doi_str_mv | 10.1007/s00792-018-1060-5 |
| format | Article |
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bop
promoter and induces bacterioopsin (Bop) expression under light and low oxygen tension. Bacterioopsin is linked to retinal to produce bacteriorhodopsin (BR), which in turn supplies energy source in
Halobacterium salinarum
. In this study, effect of Bat HTH motif–promoter DNA interaction on bacterioopsin (Bop) expression was investigated using in silico and experimental approaches. Molecular docking showed that the most stable DNA–protein complex was generated by Q661R/Q665R mutant. Based on the in silico analysis, HTH motif was mutated using site-directed mutagenesis and
Hbt. salinarum
recombinant strains were developed by introduction of mutant
bat
genes. Double positively charged amino acid substitutions (Q661R/Q665R) in second helix of HTH motif increased whereas deletion of this region decreased BR production. However, other single substitutions (Q665R and Q661H) did not change BR production. These findings represent key role of HTH motif stability for DNA binding and regulation of bacterioopsin (Bop) expression and bacteriorhodopsin (BR) production independent of environmental condition.</description><identifier>ISSN: 1431-0651</identifier><identifier>EISSN: 1433-4909</identifier><identifier>DOI: 10.1007/s00792-018-1060-5</identifier><identifier>PMID: 30350225</identifier><language>eng</language><publisher>Tokyo: Springer Japan</publisher><subject>Amino acids ; Bacteriorhodopsin ; Bacteriorhodopsins - genetics ; Bacteriorhodopsins - metabolism ; Binding ; Binding Sites ; Biochemistry ; Biomedical and Life Sciences ; Biotechnology ; Deoxyribonucleic acid ; DNA ; Energy sources ; Environmental conditions ; Gene expression ; Halobacterium salinarum ; Halobacterium salinarum - genetics ; Halobacterium salinarum - metabolism ; Industrial Microbiology - methods ; Life Sciences ; Microbial Ecology ; Microbiology ; Molecular docking ; Molecular Docking Simulation ; Mutagenesis ; Mutation, Missense ; Original Paper ; Oxygen tension ; Promoter Regions, Genetic ; Promoters ; Protein Binding ; Proteins ; Recombinants ; Retina ; Site-directed mutagenesis ; Stability ; Transcription Factors - chemistry ; Transcription Factors - genetics ; Transcription Factors - metabolism</subject><ispartof>Extremophiles : life under extreme conditions, 2019-01, Vol.23 (1), p.59-67</ispartof><rights>Springer Japan KK, part of Springer Nature 2018</rights><rights>Extremophiles is a copyright of Springer, (2018). All Rights Reserved.</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c372t-71b0680fe06c8c3eea1049b451c4f9628e65ed9d602122b1e514a02cbd1c69473</citedby><cites>FETCH-LOGICAL-c372t-71b0680fe06c8c3eea1049b451c4f9628e65ed9d602122b1e514a02cbd1c69473</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/s00792-018-1060-5$$EPDF$$P50$$Gspringer$$H</linktopdf><linktohtml>$$Uhttps://link.springer.com/10.1007/s00792-018-1060-5$$EHTML$$P50$$Gspringer$$H</linktohtml><link.rule.ids>314,780,784,27924,27925,41488,42557,51319</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/30350225$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Mirfeizollahi, Azadeh</creatorcontrib><creatorcontrib>Yakhchali, Bagher</creatorcontrib><creatorcontrib>Deldar, Ali Asghar</creatorcontrib><creatorcontrib>Karkhane, Ali Asghar</creatorcontrib><title>In silico and experimental improvement of bacteriorhodopsin production in Halobacterium salinarum R1 by increasing DNA-binding affinity of Bat through Q661R/Q665R substitutions in HTH motif</title><title>Extremophiles : life under extreme conditions</title><addtitle>Extremophiles</addtitle><addtitle>Extremophiles</addtitle><description>DNA-binding motif of bacterioopsin activator (Bat) protein is a Helix–Turn–Helix motif, which binds to
bop
promoter and induces bacterioopsin (Bop) expression under light and low oxygen tension. Bacterioopsin is linked to retinal to produce bacteriorhodopsin (BR), which in turn supplies energy source in
Halobacterium salinarum
. In this study, effect of Bat HTH motif–promoter DNA interaction on bacterioopsin (Bop) expression was investigated using in silico and experimental approaches. Molecular docking showed that the most stable DNA–protein complex was generated by Q661R/Q665R mutant. Based on the in silico analysis, HTH motif was mutated using site-directed mutagenesis and
Hbt. salinarum
recombinant strains were developed by introduction of mutant
bat
genes. Double positively charged amino acid substitutions (Q661R/Q665R) in second helix of HTH motif increased whereas deletion of this region decreased BR production. However, other single substitutions (Q665R and Q661H) did not change BR production. These findings represent key role of HTH motif stability for DNA binding and regulation of bacterioopsin (Bop) expression and bacteriorhodopsin (BR) production independent of environmental condition.</description><subject>Amino acids</subject><subject>Bacteriorhodopsin</subject><subject>Bacteriorhodopsins - genetics</subject><subject>Bacteriorhodopsins - metabolism</subject><subject>Binding</subject><subject>Binding Sites</subject><subject>Biochemistry</subject><subject>Biomedical and Life Sciences</subject><subject>Biotechnology</subject><subject>Deoxyribonucleic acid</subject><subject>DNA</subject><subject>Energy sources</subject><subject>Environmental conditions</subject><subject>Gene expression</subject><subject>Halobacterium salinarum</subject><subject>Halobacterium salinarum - genetics</subject><subject>Halobacterium salinarum - metabolism</subject><subject>Industrial Microbiology - methods</subject><subject>Life Sciences</subject><subject>Microbial Ecology</subject><subject>Microbiology</subject><subject>Molecular docking</subject><subject>Molecular Docking Simulation</subject><subject>Mutagenesis</subject><subject>Mutation, Missense</subject><subject>Original Paper</subject><subject>Oxygen tension</subject><subject>Promoter Regions, Genetic</subject><subject>Promoters</subject><subject>Protein Binding</subject><subject>Proteins</subject><subject>Recombinants</subject><subject>Retina</subject><subject>Site-directed mutagenesis</subject><subject>Stability</subject><subject>Transcription Factors - chemistry</subject><subject>Transcription Factors - genetics</subject><subject>Transcription Factors - metabolism</subject><issn>1431-0651</issn><issn>1433-4909</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2019</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><sourceid>8G5</sourceid><sourceid>ABUWG</sourceid><sourceid>AFKRA</sourceid><sourceid>AZQEC</sourceid><sourceid>BENPR</sourceid><sourceid>CCPQU</sourceid><sourceid>DWQXO</sourceid><sourceid>GNUQQ</sourceid><sourceid>GUQSH</sourceid><sourceid>M2O</sourceid><recordid>eNp1UcFu1DAQjRCIlsIHcEGWOJvOOLGzOZYW2EoViFU5W7bj7LpK7MV2EPtx_BtOd4ETl_GM3nvzNH5V9RrhHQK0l6mUjlHAFUUQQPmT6hybuqZNB93Txx4pCI5n1YuUHgCQF-B5dVZDzYExfl79uvUkudGZQJTvif25t9FN1mc1EjftY_hhl4mEgWhlcgFD3IU-7JPzpMD9bLILnpRprcZw4swTSWp0XsXSbZDoQyGYaFVRbcnN5yuqne-XXg2D8y4fFoP3KpO8i2He7shXIXBzWSrfkDTrlF2eF6P06HS_JlPIbnhZPRvUmOyr03tRffv44f56Te--fLq9vrqjpm5Zpi1qECsYLAizMrW1CqHpdMPRNEMn2MoKbvuuF8CQMY2WY6OAGd2jEV3T1hfV2-PecvH32aYsH8IcfbGURVF3HWArCguPLBNDStEOcl_-UsWDRJBLYPIYmCyBySUwyYvmzWnzrCfb_1X8SagQ2JGQCuS3Nv6z_v_W301vorc</recordid><startdate>20190101</startdate><enddate>20190101</enddate><creator>Mirfeizollahi, Azadeh</creator><creator>Yakhchali, Bagher</creator><creator>Deldar, Ali Asghar</creator><creator>Karkhane, Ali Asghar</creator><general>Springer Japan</general><general>Springer Nature B.V</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>3V.</scope><scope>7QL</scope><scope>7X7</scope><scope>7XB</scope><scope>88E</scope><scope>88I</scope><scope>8AO</scope><scope>8FE</scope><scope>8FH</scope><scope>8FI</scope><scope>8FJ</scope><scope>8FK</scope><scope>8G5</scope><scope>ABUWG</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>F1W</scope><scope>FYUFA</scope><scope>GHDGH</scope><scope>GNUQQ</scope><scope>GUQSH</scope><scope>H95</scope><scope>HCIFZ</scope><scope>K9.</scope><scope>L.G</scope><scope>LK8</scope><scope>M0S</scope><scope>M1P</scope><scope>M2O</scope><scope>M2P</scope><scope>M7N</scope><scope>M7P</scope><scope>MBDVC</scope><scope>PQEST</scope><scope>PQQKQ</scope><scope>PQUKI</scope><scope>Q9U</scope></search><sort><creationdate>20190101</creationdate><title>In silico and experimental improvement of bacteriorhodopsin production in Halobacterium salinarum R1 by increasing DNA-binding affinity of Bat through Q661R/Q665R substitutions in HTH motif</title><author>Mirfeizollahi, Azadeh ; Yakhchali, Bagher ; Deldar, Ali Asghar ; Karkhane, Ali Asghar</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c372t-71b0680fe06c8c3eea1049b451c4f9628e65ed9d602122b1e514a02cbd1c69473</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2019</creationdate><topic>Amino acids</topic><topic>Bacteriorhodopsin</topic><topic>Bacteriorhodopsins - genetics</topic><topic>Bacteriorhodopsins - metabolism</topic><topic>Binding</topic><topic>Binding Sites</topic><topic>Biochemistry</topic><topic>Biomedical and Life Sciences</topic><topic>Biotechnology</topic><topic>Deoxyribonucleic acid</topic><topic>DNA</topic><topic>Energy sources</topic><topic>Environmental conditions</topic><topic>Gene expression</topic><topic>Halobacterium salinarum</topic><topic>Halobacterium salinarum - genetics</topic><topic>Halobacterium salinarum - metabolism</topic><topic>Industrial Microbiology - methods</topic><topic>Life Sciences</topic><topic>Microbial Ecology</topic><topic>Microbiology</topic><topic>Molecular docking</topic><topic>Molecular Docking Simulation</topic><topic>Mutagenesis</topic><topic>Mutation, Missense</topic><topic>Original Paper</topic><topic>Oxygen tension</topic><topic>Promoter Regions, Genetic</topic><topic>Promoters</topic><topic>Protein Binding</topic><topic>Proteins</topic><topic>Recombinants</topic><topic>Retina</topic><topic>Site-directed mutagenesis</topic><topic>Stability</topic><topic>Transcription Factors - chemistry</topic><topic>Transcription Factors - genetics</topic><topic>Transcription Factors - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Mirfeizollahi, Azadeh</creatorcontrib><creatorcontrib>Yakhchali, Bagher</creatorcontrib><creatorcontrib>Deldar, Ali Asghar</creatorcontrib><creatorcontrib>Karkhane, Ali Asghar</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>ProQuest Central (Corporate)</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Health & Medical Collection</collection><collection>ProQuest Central (purchase pre-March 2016)</collection><collection>Medical Database (Alumni Edition)</collection><collection>Science Database (Alumni Edition)</collection><collection>ProQuest Pharma Collection</collection><collection>ProQuest SciTech Collection</collection><collection>ProQuest Natural Science Collection</collection><collection>Hospital Premium Collection</collection><collection>Hospital Premium Collection (Alumni Edition)</collection><collection>ProQuest Central (Alumni) (purchase pre-March 2016)</collection><collection>Research Library (Alumni Edition)</collection><collection>ProQuest Central (Alumni Edition)</collection><collection>ProQuest Central UK/Ireland</collection><collection>ProQuest Central Essentials</collection><collection>Biological Science Collection</collection><collection>ProQuest Central</collection><collection>Natural Science Collection</collection><collection>Environmental Sciences and Pollution Management</collection><collection>ProQuest One Community College</collection><collection>ProQuest Central Korea</collection><collection>ASFA: Aquatic Sciences and Fisheries Abstracts</collection><collection>Health Research Premium Collection</collection><collection>Health Research Premium Collection (Alumni)</collection><collection>ProQuest Central Student</collection><collection>Research Library Prep</collection><collection>Aquatic Science & Fisheries Abstracts (ASFA) 1: Biological Sciences & Living Resources</collection><collection>SciTech Premium Collection</collection><collection>ProQuest Health & Medical Complete (Alumni)</collection><collection>Aquatic Science & Fisheries Abstracts (ASFA) Professional</collection><collection>ProQuest Biological Science Collection</collection><collection>Health & Medical Collection (Alumni Edition)</collection><collection>Medical Database</collection><collection>Research Library</collection><collection>Science Database</collection><collection>Algology Mycology and Protozoology Abstracts (Microbiology C)</collection><collection>Biological Science Database</collection><collection>Research Library (Corporate)</collection><collection>ProQuest One Academic Eastern Edition (DO NOT USE)</collection><collection>ProQuest One Academic</collection><collection>ProQuest One Academic UKI Edition</collection><collection>ProQuest Central Basic</collection><jtitle>Extremophiles : life under extreme conditions</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Mirfeizollahi, Azadeh</au><au>Yakhchali, Bagher</au><au>Deldar, Ali Asghar</au><au>Karkhane, Ali Asghar</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>In silico and experimental improvement of bacteriorhodopsin production in Halobacterium salinarum R1 by increasing DNA-binding affinity of Bat through Q661R/Q665R substitutions in HTH motif</atitle><jtitle>Extremophiles : life under extreme conditions</jtitle><stitle>Extremophiles</stitle><addtitle>Extremophiles</addtitle><date>2019-01-01</date><risdate>2019</risdate><volume>23</volume><issue>1</issue><spage>59</spage><epage>67</epage><pages>59-67</pages><issn>1431-0651</issn><eissn>1433-4909</eissn><abstract>DNA-binding motif of bacterioopsin activator (Bat) protein is a Helix–Turn–Helix motif, which binds to
bop
promoter and induces bacterioopsin (Bop) expression under light and low oxygen tension. Bacterioopsin is linked to retinal to produce bacteriorhodopsin (BR), which in turn supplies energy source in
Halobacterium salinarum
. In this study, effect of Bat HTH motif–promoter DNA interaction on bacterioopsin (Bop) expression was investigated using in silico and experimental approaches. Molecular docking showed that the most stable DNA–protein complex was generated by Q661R/Q665R mutant. Based on the in silico analysis, HTH motif was mutated using site-directed mutagenesis and
Hbt. salinarum
recombinant strains were developed by introduction of mutant
bat
genes. Double positively charged amino acid substitutions (Q661R/Q665R) in second helix of HTH motif increased whereas deletion of this region decreased BR production. However, other single substitutions (Q665R and Q661H) did not change BR production. These findings represent key role of HTH motif stability for DNA binding and regulation of bacterioopsin (Bop) expression and bacteriorhodopsin (BR) production independent of environmental condition.</abstract><cop>Tokyo</cop><pub>Springer Japan</pub><pmid>30350225</pmid><doi>10.1007/s00792-018-1060-5</doi><tpages>9</tpages></addata></record> |
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| subjects | Amino acids Bacteriorhodopsin Bacteriorhodopsins - genetics Bacteriorhodopsins - metabolism Binding Binding Sites Biochemistry Biomedical and Life Sciences Biotechnology Deoxyribonucleic acid DNA Energy sources Environmental conditions Gene expression Halobacterium salinarum Halobacterium salinarum - genetics Halobacterium salinarum - metabolism Industrial Microbiology - methods Life Sciences Microbial Ecology Microbiology Molecular docking Molecular Docking Simulation Mutagenesis Mutation, Missense Original Paper Oxygen tension Promoter Regions, Genetic Promoters Protein Binding Proteins Recombinants Retina Site-directed mutagenesis Stability Transcription Factors - chemistry Transcription Factors - genetics Transcription Factors - metabolism |
| title | In silico and experimental improvement of bacteriorhodopsin production in Halobacterium salinarum R1 by increasing DNA-binding affinity of Bat through Q661R/Q665R substitutions in HTH motif |
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