Cloning, purification, crystallization and preliminary structural studies of penicillin V acylase from Bacillus subtilis
Penicillin acylase proteins are amidohydrolase enzymes that cleave penicillins at the amide bond connecting the side chain to their β‐lactam nucleus. An unannotated protein from Bacillus subtilis has been expressed in Escherichia coli, purified and confirmed to possess penicillin V acylase activity....
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| Veröffentlicht in: | Acta crystallographica. Section F, Structural biology and crystallization communications Structural biology and crystallization communications, 2005-07, Vol.61 (7), p.680-683 |
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| container_title | Acta crystallographica. Section F, Structural biology and crystallization communications |
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| creator | Rathinaswamy, Priya Pundle, Archana V. Prabhune, Asmita A. SivaRaman, Hepzibah Brannigan, James A. Dodson, Guy G. Suresh, C. G. |
| description | Penicillin acylase proteins are amidohydrolase enzymes that cleave penicillins at the amide bond connecting the side chain to their β‐lactam nucleus. An unannotated protein from Bacillus subtilis has been expressed in Escherichia coli, purified and confirmed to possess penicillin V acylase activity. The protein was crystallized using the hanging‐drop vapour‐diffusion method from a solution containing 4 M sodium formate in 100 mM Tris–HCl buffer pH 8.2. Diffraction data were collected under cryogenic conditions to a spacing of 2.5 Å. The crystals belonged to the orthorhombic space group C2221, with unit‐cell parameters a = 111.0, b = 308.0, c = 56.0 Å. The estimated Matthews coefficient was 3.23 Å3 Da−1, corresponding to 62% solvent content. The structure has been solved using molecular‐replacement methods with B. sphaericus penicillin V acylase (PDB code 2pva) as the search model. |
| doi_str_mv | 10.1107/S1744309105017987 |
| format | Article |
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G.</creator><creatorcontrib>Rathinaswamy, Priya ; Pundle, Archana V. ; Prabhune, Asmita A. ; SivaRaman, Hepzibah ; Brannigan, James A. ; Dodson, Guy G. ; Suresh, C. G.</creatorcontrib><description>Penicillin acylase proteins are amidohydrolase enzymes that cleave penicillins at the amide bond connecting the side chain to their β‐lactam nucleus. An unannotated protein from Bacillus subtilis has been expressed in Escherichia coli, purified and confirmed to possess penicillin V acylase activity. The protein was crystallized using the hanging‐drop vapour‐diffusion method from a solution containing 4 M sodium formate in 100 mM Tris–HCl buffer pH 8.2. Diffraction data were collected under cryogenic conditions to a spacing of 2.5 Å. The crystals belonged to the orthorhombic space group C2221, with unit‐cell parameters a = 111.0, b = 308.0, c = 56.0 Å. The estimated Matthews coefficient was 3.23 Å3 Da−1, corresponding to 62% solvent content. The structure has been solved using molecular‐replacement methods with B. sphaericus penicillin V acylase (PDB code 2pva) as the search model.</description><identifier>ISSN: 1744-3091</identifier><identifier>EISSN: 1744-3091</identifier><identifier>DOI: 10.1107/S1744309105017987</identifier><identifier>PMID: 16511127</identifier><language>eng</language><publisher>5 Abbey Square, Chester, Cheshire CH1 2HU, England: Munksgaard International Publishers</publisher><subject>Amino Acid Sequence ; Bacillus subtilis - enzymology ; BUFFERS ; Cloning, Molecular ; CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY ; conjugated bile-salt hydrolase ; CRYSTALLIZATION ; Crystallization Communications ; Crystallography, X-Ray - methods ; CRYSTALS ; Databases, Protein ; DIFFRACTION ; DIFFUSION ; ESCHERICHIA COLI ; FASTENING ; Formates - pharmacology ; Hydrogen-Ion Concentration ; MATHEMATICAL SOLUTIONS ; Molecular Sequence Data ; Ntn hydrolase ; Penicillin Amidase - chemistry ; penicillin V acylase ; Protein Conformation ; Recombinant Proteins - chemistry ; Sequence Homology, Amino Acid ; SODIUM ; SOLUTIONS ; SOLVENTS ; SPACE GROUPS</subject><ispartof>Acta crystallographica. 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G.</creatorcontrib><title>Cloning, purification, crystallization and preliminary structural studies of penicillin V acylase from Bacillus subtilis</title><title>Acta crystallographica. Section F, Structural biology and crystallization communications</title><addtitle>Acta Cryst. F</addtitle><description>Penicillin acylase proteins are amidohydrolase enzymes that cleave penicillins at the amide bond connecting the side chain to their β‐lactam nucleus. An unannotated protein from Bacillus subtilis has been expressed in Escherichia coli, purified and confirmed to possess penicillin V acylase activity. The protein was crystallized using the hanging‐drop vapour‐diffusion method from a solution containing 4 M sodium formate in 100 mM Tris–HCl buffer pH 8.2. Diffraction data were collected under cryogenic conditions to a spacing of 2.5 Å. The crystals belonged to the orthorhombic space group C2221, with unit‐cell parameters a = 111.0, b = 308.0, c = 56.0 Å. The estimated Matthews coefficient was 3.23 Å3 Da−1, corresponding to 62% solvent content. The structure has been solved using molecular‐replacement methods with B. sphaericus penicillin V acylase (PDB code 2pva) as the search model.</description><subject>Amino Acid Sequence</subject><subject>Bacillus subtilis - enzymology</subject><subject>BUFFERS</subject><subject>Cloning, Molecular</subject><subject>CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY</subject><subject>conjugated bile-salt hydrolase</subject><subject>CRYSTALLIZATION</subject><subject>Crystallization Communications</subject><subject>Crystallography, X-Ray - methods</subject><subject>CRYSTALS</subject><subject>Databases, Protein</subject><subject>DIFFRACTION</subject><subject>DIFFUSION</subject><subject>ESCHERICHIA COLI</subject><subject>FASTENING</subject><subject>Formates - pharmacology</subject><subject>Hydrogen-Ion Concentration</subject><subject>MATHEMATICAL SOLUTIONS</subject><subject>Molecular Sequence Data</subject><subject>Ntn hydrolase</subject><subject>Penicillin Amidase - chemistry</subject><subject>penicillin V acylase</subject><subject>Protein Conformation</subject><subject>Recombinant Proteins - chemistry</subject><subject>Sequence Homology, Amino Acid</subject><subject>SODIUM</subject><subject>SOLUTIONS</subject><subject>SOLVENTS</subject><subject>SPACE GROUPS</subject><issn>1744-3091</issn><issn>1744-3091</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2005</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkVtv1DAQhSMEohf4AbwgS0g8NeBLHG9ekMqKLlQrLioF8WTZXqcdcOxgO6XLrychq1LEA08eHX_nzIymKB4R_IwQLJ6fEVFVDDcEc0xEsxB3iv1JKift7q16rzhI6SvGjDX14n6xR2pOCKFiv7heuuDBXxyhfojQglEZgj9CJm5TVs7Bz98CUn6D-mgddOBV3KKU42DyEJUby2EDNqHQot56MDC6PPqElNk6lSxqY-jQSzXpQ0Jp0BkcpAfFvVa5ZB_u3sPi_OTVx-Xrcv1u9WZ5vC5NJTArlWBMK031RljDhairpla1UQ0RfLEQ2lpGrTasIq1iI4MFx61uMBMMa91W7LB4Mef2g-7sxlifx6FlH6Eb95BBgfz7x8OlvAhXkjScVnwKeDIHhJRBJgPZmksTvLcmS0oZr0mFR-rprk0M3websuwgGeuc8jYMSQpMGF0wMoJkBk0MKUXb3oxCsJyuKv-56uh5fHuHP47dGUegmYEf4Oz2_4ny-MsJfX_OccNGbzl7IWV7feNV8ZusBRNcfn67kvT0bPXhlK7liv0CbqC__Q</recordid><startdate>200507</startdate><enddate>200507</enddate><creator>Rathinaswamy, Priya</creator><creator>Pundle, Archana V.</creator><creator>Prabhune, Asmita A.</creator><creator>SivaRaman, Hepzibah</creator><creator>Brannigan, James A.</creator><creator>Dodson, Guy G.</creator><creator>Suresh, C. G.</creator><general>Munksgaard International Publishers</general><general>International Union of Crystallography</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><scope>OTOTI</scope><scope>5PM</scope></search><sort><creationdate>200507</creationdate><title>Cloning, purification, crystallization and preliminary structural studies of penicillin V acylase from Bacillus subtilis</title><author>Rathinaswamy, Priya ; Pundle, Archana V. ; Prabhune, Asmita A. ; SivaRaman, Hepzibah ; Brannigan, James A. ; Dodson, Guy G. ; Suresh, C. 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G.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Cloning, purification, crystallization and preliminary structural studies of penicillin V acylase from Bacillus subtilis</atitle><jtitle>Acta crystallographica. Section F, Structural biology and crystallization communications</jtitle><addtitle>Acta Cryst. F</addtitle><date>2005-07</date><risdate>2005</risdate><volume>61</volume><issue>7</issue><spage>680</spage><epage>683</epage><pages>680-683</pages><issn>1744-3091</issn><eissn>1744-3091</eissn><abstract>Penicillin acylase proteins are amidohydrolase enzymes that cleave penicillins at the amide bond connecting the side chain to their β‐lactam nucleus. An unannotated protein from Bacillus subtilis has been expressed in Escherichia coli, purified and confirmed to possess penicillin V acylase activity. The protein was crystallized using the hanging‐drop vapour‐diffusion method from a solution containing 4 M sodium formate in 100 mM Tris–HCl buffer pH 8.2. Diffraction data were collected under cryogenic conditions to a spacing of 2.5 Å. The crystals belonged to the orthorhombic space group C2221, with unit‐cell parameters a = 111.0, b = 308.0, c = 56.0 Å. The estimated Matthews coefficient was 3.23 Å3 Da−1, corresponding to 62% solvent content. The structure has been solved using molecular‐replacement methods with B. sphaericus penicillin V acylase (PDB code 2pva) as the search model.</abstract><cop>5 Abbey Square, Chester, Cheshire CH1 2HU, England</cop><pub>Munksgaard International Publishers</pub><pmid>16511127</pmid><doi>10.1107/S1744309105017987</doi><tpages>4</tpages><oa>free_for_read</oa></addata></record> |
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| subjects | Amino Acid Sequence Bacillus subtilis - enzymology BUFFERS Cloning, Molecular CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY conjugated bile-salt hydrolase CRYSTALLIZATION Crystallization Communications Crystallography, X-Ray - methods CRYSTALS Databases, Protein DIFFRACTION DIFFUSION ESCHERICHIA COLI FASTENING Formates - pharmacology Hydrogen-Ion Concentration MATHEMATICAL SOLUTIONS Molecular Sequence Data Ntn hydrolase Penicillin Amidase - chemistry penicillin V acylase Protein Conformation Recombinant Proteins - chemistry Sequence Homology, Amino Acid SODIUM SOLUTIONS SOLVENTS SPACE GROUPS |
| title | Cloning, purification, crystallization and preliminary structural studies of penicillin V acylase from Bacillus subtilis |
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