Biochemical Defects in Retina-specific Human ATP Binding Cassette Transporter Nucleotide Binding Domain 1 Mutants Associated with Macular Degeneration
The retina-specific human ABC transporter (ABCR) functions in the retinal transport system and has been implicated in several inherited visual diseases, including Stargardt disease, fundus flavimaculatus, cone-rod dystrophy, and age-related macular degeneration. We have previously described a genera...
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| Veröffentlicht in: | The Journal of biological chemistry 2002-06, Vol.277 (24), p.21759-21767 |
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| description | The retina-specific human ABC transporter (ABCR) functions in the retinal transport system and has been implicated in several
inherited visual diseases, including Stargardt disease, fundus flavimaculatus, cone-rod dystrophy, and age-related macular
degeneration. We have previously described a general ribonucleotidase activity of the first nucleotide binding domain (NBD1)
of human ABCR (Biswas, E. E. (2001) Biochemistry 40, 8181â8187). In this communication, we present a quantitative study analyzing the effects of certain disease-associated
mutations, Gly-863 â Ala, Pro-940 â Arg, and Arg-943 â Gln on the nucleotide binding, and general ribonucleotidase activities
of this domain. NBD1 proteins, harboring these mutations, were created through in vitro site-specific mutagenesis and expressed in Escherichia coli . Results of the enzyme-kinetic studies indicated that these mutations altered the ATPase and CTPase activities of NBD1. The
G863A and P940R mutations were found to have significant attenuation of the rates of nucleotide hydrolysis and binding affinities.
On the other hand, the R943Q mutation had small, but detectable reduction in its nucleotidase activity and nucleotide binding
affinity. We have measured the nucleotide binding affinities of NBD1 protein and its mutants quantitatively by fluorescence
anisotropy changes during protein binding to ethenoadenosine ATP (εATP), a fluorescent ATP analogue. We have correlated the
dissociation constant ( K
D ) and the rates of nucleotide hydrolysis ( V
max ) of NBD1 and its mutants with the available genetic data for these mutations. |
| doi_str_mv | 10.1074/jbc.M202053200 |
| format | Article |
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inherited visual diseases, including Stargardt disease, fundus flavimaculatus, cone-rod dystrophy, and age-related macular
degeneration. We have previously described a general ribonucleotidase activity of the first nucleotide binding domain (NBD1)
of human ABCR (Biswas, E. E. (2001) Biochemistry 40, 8181â8187). In this communication, we present a quantitative study analyzing the effects of certain disease-associated
mutations, Gly-863 â Ala, Pro-940 â Arg, and Arg-943 â Gln on the nucleotide binding, and general ribonucleotidase activities
of this domain. NBD1 proteins, harboring these mutations, were created through in vitro site-specific mutagenesis and expressed in Escherichia coli . Results of the enzyme-kinetic studies indicated that these mutations altered the ATPase and CTPase activities of NBD1. The
G863A and P940R mutations were found to have significant attenuation of the rates of nucleotide hydrolysis and binding affinities.
On the other hand, the R943Q mutation had small, but detectable reduction in its nucleotidase activity and nucleotide binding
affinity. We have measured the nucleotide binding affinities of NBD1 protein and its mutants quantitatively by fluorescence
anisotropy changes during protein binding to ethenoadenosine ATP (εATP), a fluorescent ATP analogue. We have correlated the
dissociation constant ( K
D ) and the rates of nucleotide hydrolysis ( V
max ) of NBD1 and its mutants with the available genetic data for these mutations.</description><identifier>ISSN: 0021-9258</identifier><identifier>EISSN: 1083-351X</identifier><identifier>DOI: 10.1074/jbc.M202053200</identifier><identifier>PMID: 11919200</identifier><language>eng</language><publisher>United States: American Society for Biochemistry and Molecular Biology</publisher><subject>Adenosine Triphosphatases - chemistry ; Adenosine Triphosphate - metabolism ; Amino Acid Sequence ; Anisotropy ; Arginine - chemistry ; ATP-Binding Cassette Transporters - chemistry ; ATP-Binding Cassette Transporters - metabolism ; Cloning, Molecular ; Databases as Topic ; Dose-Response Relationship, Drug ; Electrophoresis, Polyacrylamide Gel ; Escherichia coli - metabolism ; fluorescence anisotropy ; Glycine - chemistry ; Humans ; Hydrolysis ; Kinetics ; macular degeneration ; Macular Degeneration - metabolism ; Models, Molecular ; Molecular Sequence Data ; Mutagenesis, Site-Directed ; Mutation ; NBD1 protein ; Phenotype ; Proline - chemistry ; Protein Binding ; Protein Structure, Tertiary ; Pyrophosphatases - chemistry ; Recombinant Proteins - chemistry ; Recombinant Proteins - metabolism ; Retina - metabolism ; ribonucleotidase ; Sequence Homology, Amino Acid ; Spectrometry, Fluorescence ; Thermodynamics ; Time Factors</subject><ispartof>The Journal of biological chemistry, 2002-06, Vol.277 (24), p.21759-21767</ispartof><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c391t-aadd54ddf0fd892ec5bfb43972824ed262196ae9fe46ba918531848f188e9203</citedby><cites>FETCH-LOGICAL-c391t-aadd54ddf0fd892ec5bfb43972824ed262196ae9fe46ba918531848f188e9203</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,776,780,27903,27904</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/11919200$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Suárez, Tatiana</creatorcontrib><creatorcontrib>Biswas, Subhasis B</creatorcontrib><creatorcontrib>Biswas, Esther E</creatorcontrib><title>Biochemical Defects in Retina-specific Human ATP Binding Cassette Transporter Nucleotide Binding Domain 1 Mutants Associated with Macular Degeneration</title><title>The Journal of biological chemistry</title><addtitle>J Biol Chem</addtitle><description>The retina-specific human ABC transporter (ABCR) functions in the retinal transport system and has been implicated in several
inherited visual diseases, including Stargardt disease, fundus flavimaculatus, cone-rod dystrophy, and age-related macular
degeneration. We have previously described a general ribonucleotidase activity of the first nucleotide binding domain (NBD1)
of human ABCR (Biswas, E. E. (2001) Biochemistry 40, 8181â8187). In this communication, we present a quantitative study analyzing the effects of certain disease-associated
mutations, Gly-863 â Ala, Pro-940 â Arg, and Arg-943 â Gln on the nucleotide binding, and general ribonucleotidase activities
of this domain. NBD1 proteins, harboring these mutations, were created through in vitro site-specific mutagenesis and expressed in Escherichia coli . Results of the enzyme-kinetic studies indicated that these mutations altered the ATPase and CTPase activities of NBD1. The
G863A and P940R mutations were found to have significant attenuation of the rates of nucleotide hydrolysis and binding affinities.
On the other hand, the R943Q mutation had small, but detectable reduction in its nucleotidase activity and nucleotide binding
affinity. We have measured the nucleotide binding affinities of NBD1 protein and its mutants quantitatively by fluorescence
anisotropy changes during protein binding to ethenoadenosine ATP (εATP), a fluorescent ATP analogue. We have correlated the
dissociation constant ( K
D ) and the rates of nucleotide hydrolysis ( V
max ) of NBD1 and its mutants with the available genetic data for these mutations.</description><subject>Adenosine Triphosphatases - chemistry</subject><subject>Adenosine Triphosphate - metabolism</subject><subject>Amino Acid Sequence</subject><subject>Anisotropy</subject><subject>Arginine - chemistry</subject><subject>ATP-Binding Cassette Transporters - chemistry</subject><subject>ATP-Binding Cassette Transporters - metabolism</subject><subject>Cloning, Molecular</subject><subject>Databases as Topic</subject><subject>Dose-Response Relationship, Drug</subject><subject>Electrophoresis, Polyacrylamide Gel</subject><subject>Escherichia coli - metabolism</subject><subject>fluorescence anisotropy</subject><subject>Glycine - chemistry</subject><subject>Humans</subject><subject>Hydrolysis</subject><subject>Kinetics</subject><subject>macular degeneration</subject><subject>Macular Degeneration - metabolism</subject><subject>Models, Molecular</subject><subject>Molecular Sequence Data</subject><subject>Mutagenesis, Site-Directed</subject><subject>Mutation</subject><subject>NBD1 protein</subject><subject>Phenotype</subject><subject>Proline - chemistry</subject><subject>Protein Binding</subject><subject>Protein Structure, Tertiary</subject><subject>Pyrophosphatases - chemistry</subject><subject>Recombinant Proteins - chemistry</subject><subject>Recombinant Proteins - metabolism</subject><subject>Retina - metabolism</subject><subject>ribonucleotidase</subject><subject>Sequence Homology, Amino Acid</subject><subject>Spectrometry, Fluorescence</subject><subject>Thermodynamics</subject><subject>Time Factors</subject><issn>0021-9258</issn><issn>1083-351X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2002</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNpFkU1vEzEQhi0EomnhyhH5gHrb4K_N2sc0pRSpAYRy4GZ57dnE1a4dbK8q_gi_F6NEdC5zeeYZzbwIvaNkSUknPj72drllhJGWM0JeoAUlkje8pT9fogUhjDaKtfICXeb8SGoJRV-jC0oVVZVfoD83PtoDTN6aEd_CALZk7AP-AcUH0-QjWD94i-_nyQS83n3HNz44H_Z4Y3KGUgDvkgn5GFOBhL_OdoRYvIP_3G2cTBVSvJ2LCdW-zjlabwo4_OTLAW-NnUeT6vY9BEim-BjeoFeDGTO8PfcrtLv7tNvcNw_fPn_ZrB8ayxUtjTHOtcK5gQxOKga27YdecNUxyQQ4tmJUrQyoAcSqN4rKllMp5EClhHo_v0LXJ-0xxV8z5KInny2MowkQ56yp5IIo1VZweQJtijknGPQx-cmk35oS_S8IXYPQz0HUgfdn89xP4J7x8-cr8OEEHPz-8OQT6P6UhGZdp5nQjHat4n8BlRiR1g</recordid><startdate>20020614</startdate><enddate>20020614</enddate><creator>Suárez, Tatiana</creator><creator>Biswas, Subhasis B</creator><creator>Biswas, Esther E</creator><general>American Society for Biochemistry and Molecular Biology</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>8FD</scope><scope>FR3</scope><scope>P64</scope><scope>RC3</scope></search><sort><creationdate>20020614</creationdate><title>Biochemical Defects in Retina-specific Human ATP Binding Cassette Transporter Nucleotide Binding Domain 1 Mutants Associated with Macular Degeneration</title><author>Suárez, Tatiana ; Biswas, Subhasis B ; Biswas, Esther E</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c391t-aadd54ddf0fd892ec5bfb43972824ed262196ae9fe46ba918531848f188e9203</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2002</creationdate><topic>Adenosine Triphosphatases - chemistry</topic><topic>Adenosine Triphosphate - metabolism</topic><topic>Amino Acid Sequence</topic><topic>Anisotropy</topic><topic>Arginine - chemistry</topic><topic>ATP-Binding Cassette Transporters - chemistry</topic><topic>ATP-Binding Cassette Transporters - metabolism</topic><topic>Cloning, Molecular</topic><topic>Databases as Topic</topic><topic>Dose-Response Relationship, Drug</topic><topic>Electrophoresis, Polyacrylamide Gel</topic><topic>Escherichia coli - metabolism</topic><topic>fluorescence anisotropy</topic><topic>Glycine - chemistry</topic><topic>Humans</topic><topic>Hydrolysis</topic><topic>Kinetics</topic><topic>macular degeneration</topic><topic>Macular Degeneration - metabolism</topic><topic>Models, Molecular</topic><topic>Molecular Sequence Data</topic><topic>Mutagenesis, Site-Directed</topic><topic>Mutation</topic><topic>NBD1 protein</topic><topic>Phenotype</topic><topic>Proline - chemistry</topic><topic>Protein Binding</topic><topic>Protein Structure, Tertiary</topic><topic>Pyrophosphatases - chemistry</topic><topic>Recombinant Proteins - chemistry</topic><topic>Recombinant Proteins - metabolism</topic><topic>Retina - metabolism</topic><topic>ribonucleotidase</topic><topic>Sequence Homology, Amino Acid</topic><topic>Spectrometry, Fluorescence</topic><topic>Thermodynamics</topic><topic>Time Factors</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Suárez, Tatiana</creatorcontrib><creatorcontrib>Biswas, Subhasis B</creatorcontrib><creatorcontrib>Biswas, Esther E</creatorcontrib><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>Biotechnology and BioEngineering Abstracts</collection><collection>Genetics Abstracts</collection><jtitle>The Journal of biological chemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Suárez, Tatiana</au><au>Biswas, Subhasis B</au><au>Biswas, Esther E</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Biochemical Defects in Retina-specific Human ATP Binding Cassette Transporter Nucleotide Binding Domain 1 Mutants Associated with Macular Degeneration</atitle><jtitle>The Journal of biological chemistry</jtitle><addtitle>J Biol Chem</addtitle><date>2002-06-14</date><risdate>2002</risdate><volume>277</volume><issue>24</issue><spage>21759</spage><epage>21767</epage><pages>21759-21767</pages><issn>0021-9258</issn><eissn>1083-351X</eissn><abstract>The retina-specific human ABC transporter (ABCR) functions in the retinal transport system and has been implicated in several
inherited visual diseases, including Stargardt disease, fundus flavimaculatus, cone-rod dystrophy, and age-related macular
degeneration. We have previously described a general ribonucleotidase activity of the first nucleotide binding domain (NBD1)
of human ABCR (Biswas, E. E. (2001) Biochemistry 40, 8181â8187). In this communication, we present a quantitative study analyzing the effects of certain disease-associated
mutations, Gly-863 â Ala, Pro-940 â Arg, and Arg-943 â Gln on the nucleotide binding, and general ribonucleotidase activities
of this domain. NBD1 proteins, harboring these mutations, were created through in vitro site-specific mutagenesis and expressed in Escherichia coli . Results of the enzyme-kinetic studies indicated that these mutations altered the ATPase and CTPase activities of NBD1. The
G863A and P940R mutations were found to have significant attenuation of the rates of nucleotide hydrolysis and binding affinities.
On the other hand, the R943Q mutation had small, but detectable reduction in its nucleotidase activity and nucleotide binding
affinity. We have measured the nucleotide binding affinities of NBD1 protein and its mutants quantitatively by fluorescence
anisotropy changes during protein binding to ethenoadenosine ATP (εATP), a fluorescent ATP analogue. We have correlated the
dissociation constant ( K
D ) and the rates of nucleotide hydrolysis ( V
max ) of NBD1 and its mutants with the available genetic data for these mutations.</abstract><cop>United States</cop><pub>American Society for Biochemistry and Molecular Biology</pub><pmid>11919200</pmid><doi>10.1074/jbc.M202053200</doi><tpages>9</tpages><oa>free_for_read</oa></addata></record> |
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| issn | 0021-9258 1083-351X |
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| source | MEDLINE; EZB-FREE-00999 freely available EZB journals; Alma/SFX Local Collection |
| subjects | Adenosine Triphosphatases - chemistry Adenosine Triphosphate - metabolism Amino Acid Sequence Anisotropy Arginine - chemistry ATP-Binding Cassette Transporters - chemistry ATP-Binding Cassette Transporters - metabolism Cloning, Molecular Databases as Topic Dose-Response Relationship, Drug Electrophoresis, Polyacrylamide Gel Escherichia coli - metabolism fluorescence anisotropy Glycine - chemistry Humans Hydrolysis Kinetics macular degeneration Macular Degeneration - metabolism Models, Molecular Molecular Sequence Data Mutagenesis, Site-Directed Mutation NBD1 protein Phenotype Proline - chemistry Protein Binding Protein Structure, Tertiary Pyrophosphatases - chemistry Recombinant Proteins - chemistry Recombinant Proteins - metabolism Retina - metabolism ribonucleotidase Sequence Homology, Amino Acid Spectrometry, Fluorescence Thermodynamics Time Factors |
| title | Biochemical Defects in Retina-specific Human ATP Binding Cassette Transporter Nucleotide Binding Domain 1 Mutants Associated with Macular Degeneration |
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