X-ray studies of recombinant anti-testosterone Fab fragments: the use of PEG 3350 in crystallization
Recombinant anti‐testosterone wild‐type Fab fragment and mutant Fab fragments with high binding selectivity developed by protein engineering have been crystallized with and without ligands. Crystals of these Fab fragments were obtained by the vapour‐diffusion technique at room temperature using solu...
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| Veröffentlicht in: | Acta crystallographica. Section D, Biological crystallography. Biological crystallography., 2000-02, Vol.56 (2), p.218-221 |
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| container_title | Acta crystallographica. Section D, Biological crystallography. |
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| creator | Valjakka, Jarkko Hemminki, Ari Teerinen, Tuija Takkinen, Kristiina Rouvinen, Juha |
| description | Recombinant anti‐testosterone wild‐type Fab fragment and mutant Fab fragments with high binding selectivity developed by protein engineering have been crystallized with and without ligands. Crystals of these Fab fragments were obtained by the vapour‐diffusion technique at room temperature using solutions of PEG 3350 with various biological buffers and with a wide pH range. So far, five data sets have been collected from crystals of three Fab–antigen complexes and from two uncomplexed Fab fragments, with resolutions ranging from 2.10 to 3.1 Å. Crystallization conditions for Fab fragments were found by using modifications of the low ionic strength PEG 3350 series. Suitable concentrations of PEG 400, MPD and glycerol solutions for use as cryoprotectants in PEG 3350 solutions have been determined. One useful observation was that PEG 3350 is able to work alone as a cryoprotectant. The screening protocol used requires a smaller amount of protein material to achieve auspicious pre‐crystals than previously. Results support the claim that PEG 3350 is more suitable for the crystallization of Fab fragments than higher molecular weight PEGs. |
| doi_str_mv | 10.1107/S0907444999016224 |
| format | Article |
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Section D, Biological crystallography.</title><addtitle>Acta Cryst. D</addtitle><description>Recombinant anti‐testosterone wild‐type Fab fragment and mutant Fab fragments with high binding selectivity developed by protein engineering have been crystallized with and without ligands. Crystals of these Fab fragments were obtained by the vapour‐diffusion technique at room temperature using solutions of PEG 3350 with various biological buffers and with a wide pH range. So far, five data sets have been collected from crystals of three Fab–antigen complexes and from two uncomplexed Fab fragments, with resolutions ranging from 2.10 to 3.1 Å. Crystallization conditions for Fab fragments were found by using modifications of the low ionic strength PEG 3350 series. Suitable concentrations of PEG 400, MPD and glycerol solutions for use as cryoprotectants in PEG 3350 solutions have been determined. One useful observation was that PEG 3350 is able to work alone as a cryoprotectant. The screening protocol used requires a smaller amount of protein material to achieve auspicious pre‐crystals than previously. Results support the claim that PEG 3350 is more suitable for the crystallization of Fab fragments than higher molecular weight PEGs.</description><subject>Buffers</subject><subject>Chemical Precipitation</subject><subject>Cold Temperature</subject><subject>Crystallization</subject><subject>Dehydroepiandrosterone Sulfate - chemistry</subject><subject>Diffusion</subject><subject>Immunoglobulin Fragments - chemistry</subject><subject>Osmolar Concentration</subject><subject>Polyethylene Glycols - chemistry</subject><subject>recombinant Fab fragments</subject><subject>Recombinant Proteins - chemistry</subject><subject>testosterone</subject><subject>Testosterone - chemistry</subject><subject>Testosterone - immunology</subject><subject>Volatilization</subject><issn>1399-0047</issn><issn>0907-4449</issn><issn>1399-0047</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2000</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkMtuFDEQRS0EIiHwAWyQV-waqmynK2YXJZMBaRhA4RFWlu12g6EfwXYLhq-nRx2hSCwoySov7rkqHcYeIzxDBHp-CRpIKaW1BqyFUHfYIUqtKwBFd2_9D9iDnL8BgBCS7rMDhHoexEPWXFXJ7nguUxND5mPLU_Bj7-Jgh8LnF6sSchlzCWkcAr-wjrfJfunDUPILXr4GPuWw596u1lzKY-Bx4D7tcrFdF3_bEsfhIbvX2i6HRzf7iH24WL0_e1lt3qxfnZ1uKi-0xsq1wTtFyhJ5sJoInHNaulbU3jbWhxPEGgVZoYNER857EqBsowkUNrU8Yk-X3us0_pjms00fsw9dZ4cwTtnQ7EujFHMQl6BPY84ptOY6xd6mnUEwe7XmH7Uz8-SmfHJ9aG4Ri8s5cLIEfsYu7P7faE4_n1-uQOg9Wi1onD3_-ova9N3UJOnYfNquzXb77vUVbaT5KP8AzOuTRA</recordid><startdate>200002</startdate><enddate>200002</enddate><creator>Valjakka, Jarkko</creator><creator>Hemminki, Ari</creator><creator>Teerinen, Tuija</creator><creator>Takkinen, Kristiina</creator><creator>Rouvinen, Juha</creator><general>Munksgaard International Publishers</general><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>7X8</scope></search><sort><creationdate>200002</creationdate><title>X-ray studies of recombinant anti-testosterone Fab fragments: the use of PEG 3350 in crystallization</title><author>Valjakka, Jarkko ; Hemminki, Ari ; Teerinen, Tuija ; Takkinen, Kristiina ; Rouvinen, Juha</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c2991-bfecb474a77c0a9770bbb93bf26cadace8116127a29e31b7bcc7204ad97041d63</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2000</creationdate><topic>Buffers</topic><topic>Chemical Precipitation</topic><topic>Cold Temperature</topic><topic>Crystallization</topic><topic>Dehydroepiandrosterone Sulfate - chemistry</topic><topic>Diffusion</topic><topic>Immunoglobulin Fragments - chemistry</topic><topic>Osmolar Concentration</topic><topic>Polyethylene Glycols - chemistry</topic><topic>recombinant Fab fragments</topic><topic>Recombinant Proteins - chemistry</topic><topic>testosterone</topic><topic>Testosterone - chemistry</topic><topic>Testosterone - immunology</topic><topic>Volatilization</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Valjakka, Jarkko</creatorcontrib><creatorcontrib>Hemminki, Ari</creatorcontrib><creatorcontrib>Teerinen, Tuija</creatorcontrib><creatorcontrib>Takkinen, Kristiina</creatorcontrib><creatorcontrib>Rouvinen, Juha</creatorcontrib><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>MEDLINE - Academic</collection><jtitle>Acta crystallographica. Section D, Biological crystallography.</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Valjakka, Jarkko</au><au>Hemminki, Ari</au><au>Teerinen, Tuija</au><au>Takkinen, Kristiina</au><au>Rouvinen, Juha</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>X-ray studies of recombinant anti-testosterone Fab fragments: the use of PEG 3350 in crystallization</atitle><jtitle>Acta crystallographica. Section D, Biological crystallography.</jtitle><addtitle>Acta Cryst. D</addtitle><date>2000-02</date><risdate>2000</risdate><volume>56</volume><issue>2</issue><spage>218</spage><epage>221</epage><pages>218-221</pages><issn>1399-0047</issn><issn>0907-4449</issn><eissn>1399-0047</eissn><abstract>Recombinant anti‐testosterone wild‐type Fab fragment and mutant Fab fragments with high binding selectivity developed by protein engineering have been crystallized with and without ligands. Crystals of these Fab fragments were obtained by the vapour‐diffusion technique at room temperature using solutions of PEG 3350 with various biological buffers and with a wide pH range. So far, five data sets have been collected from crystals of three Fab–antigen complexes and from two uncomplexed Fab fragments, with resolutions ranging from 2.10 to 3.1 Å. Crystallization conditions for Fab fragments were found by using modifications of the low ionic strength PEG 3350 series. Suitable concentrations of PEG 400, MPD and glycerol solutions for use as cryoprotectants in PEG 3350 solutions have been determined. One useful observation was that PEG 3350 is able to work alone as a cryoprotectant. The screening protocol used requires a smaller amount of protein material to achieve auspicious pre‐crystals than previously. 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| ispartof | Acta crystallographica. Section D, Biological crystallography., 2000-02, Vol.56 (2), p.218-221 |
| issn | 1399-0047 0907-4449 1399-0047 |
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| source | MEDLINE; Crystallography Journals Online; Wiley Online Library Journals Frontfile Complete; Alma/SFX Local Collection |
| subjects | Buffers Chemical Precipitation Cold Temperature Crystallization Dehydroepiandrosterone Sulfate - chemistry Diffusion Immunoglobulin Fragments - chemistry Osmolar Concentration Polyethylene Glycols - chemistry recombinant Fab fragments Recombinant Proteins - chemistry testosterone Testosterone - chemistry Testosterone - immunology Volatilization |
| title | X-ray studies of recombinant anti-testosterone Fab fragments: the use of PEG 3350 in crystallization |
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