Switching Antibody Specificity through Minimal Mutation
Antibody 1E9, which was elicited with a hexachloronorbornene derivative and catalyzes the Diels–Alder reaction between tetrachlorothiophene dioxide and N-ethylmaleimide with high efficiency, was successfully reengineered to bind a range of structurally diverse steroids with nanomolar affinities. Rem...
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| Veröffentlicht in: | Journal of molecular biology 2008-04, Vol.377 (4), p.993-1001 |
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| container_title | Journal of molecular biology |
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| creator | Piatesi, Andrea Aldag, Caroline Hilvert, Donald |
| description | Antibody 1E9, which was elicited with a hexachloronorbornene derivative and catalyzes the Diels–Alder reaction between tetrachlorothiophene dioxide and N-ethylmaleimide with high efficiency, was successfully reengineered to bind a range of structurally diverse steroids with nanomolar affinities. Remarkably, two mutations (LeuH47Trp/ArgH100Trp) out of 36 total sequence differences suffice to switch the selectivity of 1E9 to that of the progesterone-binding antibody DB3. In contrast to the double mutant, which tightly binds multiple steroids with differently configured A–B ring junctions, the individual LeuH47Trp and ArgH100Trp single mutants both exhibit significantly greater specificity than DB3, preferentially binding 5α-pregnan-3β-ol-20-one (Kd≈5 nM) over other steroids. These findings illustrate how easily differently shaped binding pockets can be created through subtle changes to the same primordial germ line template. |
| doi_str_mv | 10.1016/j.jmb.2008.01.069 |
| format | Article |
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Remarkably, two mutations (LeuH47Trp/ArgH100Trp) out of 36 total sequence differences suffice to switch the selectivity of 1E9 to that of the progesterone-binding antibody DB3. In contrast to the double mutant, which tightly binds multiple steroids with differently configured A–B ring junctions, the individual LeuH47Trp and ArgH100Trp single mutants both exhibit significantly greater specificity than DB3, preferentially binding 5α-pregnan-3β-ol-20-one (Kd≈5 nM) over other steroids. 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Remarkably, two mutations (LeuH47Trp/ArgH100Trp) out of 36 total sequence differences suffice to switch the selectivity of 1E9 to that of the progesterone-binding antibody DB3. In contrast to the double mutant, which tightly binds multiple steroids with differently configured A–B ring junctions, the individual LeuH47Trp and ArgH100Trp single mutants both exhibit significantly greater specificity than DB3, preferentially binding 5α-pregnan-3β-ol-20-one (Kd≈5 nM) over other steroids. These findings illustrate how easily differently shaped binding pockets can be created through subtle changes to the same primordial germ line template.</description><subject>Amino Acid Sequence</subject><subject>Amino Acid Substitution</subject><subject>Antibodies, Monoclonal - genetics</subject><subject>Antibodies, Monoclonal - metabolism</subject><subject>Antibody Specificity</subject><subject>Binding Sites, Antibody - genetics</subject><subject>complementarity</subject><subject>Cross Reactions - genetics</subject><subject>cross-reactivity</subject><subject>immunoglobulin</subject><subject>Models, Biological</subject><subject>Models, Molecular</subject><subject>molecular recognition</subject><subject>Molecular Sequence Data</subject><subject>Mutagenesis, Site-Directed - methods</subject><subject>Mutant Proteins - metabolism</subject><subject>Mutation - physiology</subject><subject>Progesterone - immunology</subject><subject>Progesterone - metabolism</subject><subject>Sequence Homology, Amino Acid</subject><subject>steroid</subject><subject>Steroids - immunology</subject><subject>Steroids - metabolism</subject><issn>0022-2836</issn><issn>1089-8638</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2008</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp9kDtPwzAUhS0EoqXwA1hQJraEaztxHDFVFS-pFUO7W47ttI7yKLYD6r8nVSuxMd3lO0fnfgjdY0gwYPZUJ3VbJgSAJ4ATYMUFmmLgRcwZ5ZdoCkBITDhlE3TjfQ0AGU35NZpgToosL8gU5esfG9TOdtto3gVb9voQrfdG2coqGw5R2Ll-2O6ile1sK5toNQQZbN_doqtKNt7cne8MbV5fNov3ePn59rGYL2OVpjzEJaaKaaZzU5SYUyxJoSWTWpOSVVQCxynNTFmmyuSYyjzL0gpLUJhzldOKztDjqXbv-q_B-CBa65VpGtmZfvCCQDHmOB9BfAKV6713phJ7Nw52B4FBHGWJWoyyxFGWACxGWWPm4Vw-lK3Rf4mznRF4PgFm_PDbGie8sqZTRltnVBC6t__U_wK7WnpE</recordid><startdate>20080404</startdate><enddate>20080404</enddate><creator>Piatesi, Andrea</creator><creator>Aldag, Caroline</creator><creator>Hilvert, Donald</creator><general>Elsevier Ltd</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>20080404</creationdate><title>Switching Antibody Specificity through Minimal Mutation</title><author>Piatesi, Andrea ; Aldag, Caroline ; Hilvert, Donald</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c448t-b13c6d6d7e9b1831a29da6add2b6f3a081435ebb4ce713a7554f1a0c188c73f3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2008</creationdate><topic>Amino Acid Sequence</topic><topic>Amino Acid Substitution</topic><topic>Antibodies, Monoclonal - genetics</topic><topic>Antibodies, Monoclonal - metabolism</topic><topic>Antibody Specificity</topic><topic>Binding Sites, Antibody - genetics</topic><topic>complementarity</topic><topic>Cross Reactions - genetics</topic><topic>cross-reactivity</topic><topic>immunoglobulin</topic><topic>Models, Biological</topic><topic>Models, Molecular</topic><topic>molecular recognition</topic><topic>Molecular Sequence Data</topic><topic>Mutagenesis, Site-Directed - methods</topic><topic>Mutant Proteins - metabolism</topic><topic>Mutation - physiology</topic><topic>Progesterone - immunology</topic><topic>Progesterone - metabolism</topic><topic>Sequence Homology, Amino Acid</topic><topic>steroid</topic><topic>Steroids - immunology</topic><topic>Steroids - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Piatesi, Andrea</creatorcontrib><creatorcontrib>Aldag, Caroline</creatorcontrib><creatorcontrib>Hilvert, Donald</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>Journal of molecular biology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Piatesi, Andrea</au><au>Aldag, Caroline</au><au>Hilvert, Donald</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Switching Antibody Specificity through Minimal Mutation</atitle><jtitle>Journal of molecular biology</jtitle><addtitle>J Mol Biol</addtitle><date>2008-04-04</date><risdate>2008</risdate><volume>377</volume><issue>4</issue><spage>993</spage><epage>1001</epage><pages>993-1001</pages><issn>0022-2836</issn><eissn>1089-8638</eissn><abstract>Antibody 1E9, which was elicited with a hexachloronorbornene derivative and catalyzes the Diels–Alder reaction between tetrachlorothiophene dioxide and N-ethylmaleimide with high efficiency, was successfully reengineered to bind a range of structurally diverse steroids with nanomolar affinities. Remarkably, two mutations (LeuH47Trp/ArgH100Trp) out of 36 total sequence differences suffice to switch the selectivity of 1E9 to that of the progesterone-binding antibody DB3. In contrast to the double mutant, which tightly binds multiple steroids with differently configured A–B ring junctions, the individual LeuH47Trp and ArgH100Trp single mutants both exhibit significantly greater specificity than DB3, preferentially binding 5α-pregnan-3β-ol-20-one (Kd≈5 nM) over other steroids. These findings illustrate how easily differently shaped binding pockets can be created through subtle changes to the same primordial germ line template.</abstract><cop>England</cop><pub>Elsevier Ltd</pub><pmid>18295792</pmid><doi>10.1016/j.jmb.2008.01.069</doi><tpages>9</tpages></addata></record> |
| fulltext | fulltext |
| identifier | ISSN: 0022-2836 |
| ispartof | Journal of molecular biology, 2008-04, Vol.377 (4), p.993-1001 |
| issn | 0022-2836 1089-8638 |
| language | eng |
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| subjects | Amino Acid Sequence Amino Acid Substitution Antibodies, Monoclonal - genetics Antibodies, Monoclonal - metabolism Antibody Specificity Binding Sites, Antibody - genetics complementarity Cross Reactions - genetics cross-reactivity immunoglobulin Models, Biological Models, Molecular molecular recognition Molecular Sequence Data Mutagenesis, Site-Directed - methods Mutant Proteins - metabolism Mutation - physiology Progesterone - immunology Progesterone - metabolism Sequence Homology, Amino Acid steroid Steroids - immunology Steroids - metabolism |
| title | Switching Antibody Specificity through Minimal Mutation |
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