Investigations on three genes in Ralstonia eutropha H16 encoding putative cyanophycin metabolizing enzymes.
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| Title: | Investigations on three genes in Ralstonia eutropha H16 encoding putative cyanophycin metabolizing enzymes. |
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| Authors: | Adames, Katja1, Euting, Karina1, Bröker, Anna1, Steinbüchel, Alexander steinbu@uni-muenster.de |
| Source: | Applied Microbiology & Biotechnology. Apr2013, Vol. 97 Issue 8, p3579-3591. 13p. |
| Subjects: | Ralstonia eutropha, Amino acids, Cyanophycin, Ligases, Enzymes |
| Abstract: | The genome sequence of the facultative chemolithoautotrophic bacterium Ralstonia eutropha H16 exhibited two coding sequences with high homologies to cyanophycin synthetases (CphA) as well as one gene coding for a putative cyanophycinase (CphB). To investigate whether or not the genes cphA (H16_A0774), cphA′ (H16_A0775) and cphB (H16_B1013) encode active cyanophycin (CGP) metabolism proteins, several functional analyses were performed. Extensive in silico analysis revealed that all characteristic motifs are conserved within CphA, whereas CphA′ misses a large part of the so-called J-loop present in other active cyanophycin synthetases. Although transcription of both genes was demonstrated by RT-PCR, and heterologously expressed cphA genes led to light-scattering inclusions in recombinant cells of Escherichia coli, no CGP could be isolated from the cells or detected by HPLC analysis. For all enzyme assay experiments carried out, significant enzyme activities were determined for CphA and CphA′ in recombinant E. coli cells if crude cell extracts were applied. Homologous expression of cphA genes in cells of R. eutropha H16∆ phaC1 did not result in the formation of light-scattering inclusions, and no CGP could be isolated from the cells or detected by HPLC analysis. No transcription of cphB encoding a putative cyanophycinase could be detected by RT-PCR analysis and no overexpression was achieved in several strains of E. coli. Furthermore, no enzyme activity was detected by using CGP overlay agar plates. [ABSTRACT FROM AUTHOR] |
| Copyright of Applied Microbiology & Biotechnology is the property of Springer Nature and its content may not be copied or emailed to multiple sites without the copyright holder's express written permission. Additionally, content may not be used with any artificial intelligence tools or machine learning technologies. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.) | |
| Database: | Engineering Source |
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| Items | – Name: Title Label: Title Group: Ti Data: Investigations on three genes in Ralstonia eutropha H16 encoding putative cyanophycin metabolizing enzymes. – Name: Author Label: Authors Group: Au Data: <searchLink fieldCode="AR" term="%22Adames%2C+Katja%22">Adames, Katja</searchLink><relatesTo>1</relatesTo><br /><searchLink fieldCode="AR" term="%22Euting%2C+Karina%22">Euting, Karina</searchLink><relatesTo>1</relatesTo><br /><searchLink fieldCode="AR" term="%22Bröker%2C+Anna%22">Bröker, Anna</searchLink><relatesTo>1</relatesTo><br /><searchLink fieldCode="AR" term="%22Steinbüchel%2C+Alexander%22">Steinbüchel, Alexander</searchLink><i> steinbu@uni-muenster.de</i> – Name: TitleSource Label: Source Group: Src Data: <searchLink fieldCode="JN" term="%22Applied+Microbiology+%26+Biotechnology%22">Applied Microbiology & Biotechnology</searchLink>. Apr2013, Vol. 97 Issue 8, p3579-3591. 13p. – Name: Subject Label: Subjects Group: Su Data: <searchLink fieldCode="DE" term="%22Ralstonia+eutropha%22">Ralstonia eutropha</searchLink><br /><searchLink fieldCode="DE" term="%22Amino+acids%22">Amino acids</searchLink><br /><searchLink fieldCode="DE" term="%22Cyanophycin%22">Cyanophycin</searchLink><br /><searchLink fieldCode="DE" term="%22Ligases%22">Ligases</searchLink><br /><searchLink fieldCode="DE" term="%22Enzymes%22">Enzymes</searchLink> – Name: Abstract Label: Abstract Group: Ab Data: The genome sequence of the facultative chemolithoautotrophic bacterium Ralstonia eutropha H16 exhibited two coding sequences with high homologies to cyanophycin synthetases (CphA) as well as one gene coding for a putative cyanophycinase (CphB). To investigate whether or not the genes cphA (H16_A0774), cphA′ (H16_A0775) and cphB (H16_B1013) encode active cyanophycin (CGP) metabolism proteins, several functional analyses were performed. Extensive in silico analysis revealed that all characteristic motifs are conserved within CphA, whereas CphA′ misses a large part of the so-called J-loop present in other active cyanophycin synthetases. Although transcription of both genes was demonstrated by RT-PCR, and heterologously expressed cphA genes led to light-scattering inclusions in recombinant cells of Escherichia coli, no CGP could be isolated from the cells or detected by HPLC analysis. For all enzyme assay experiments carried out, significant enzyme activities were determined for CphA and CphA′ in recombinant E. coli cells if crude cell extracts were applied. Homologous expression of cphA genes in cells of R. eutropha H16∆ phaC1 did not result in the formation of light-scattering inclusions, and no CGP could be isolated from the cells or detected by HPLC analysis. No transcription of cphB encoding a putative cyanophycinase could be detected by RT-PCR analysis and no overexpression was achieved in several strains of E. coli. Furthermore, no enzyme activity was detected by using CGP overlay agar plates. [ABSTRACT FROM AUTHOR] – Name: AbstractSuppliedCopyright Label: Group: Ab Data: <i>Copyright of Applied Microbiology & Biotechnology is the property of Springer Nature and its content may not be copied or emailed to multiple sites without the copyright holder's express written permission. Additionally, content may not be used with any artificial intelligence tools or machine learning technologies. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract.</i> (Copyright applies to all Abstracts.) |
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| RecordInfo | BibRecord: BibEntity: Identifiers: – Type: doi Value: 10.1007/s00253-012-4599-6 Languages: – Code: eng Text: English PhysicalDescription: Pagination: PageCount: 13 StartPage: 3579 Subjects: – SubjectFull: Ralstonia eutropha Type: general – SubjectFull: Amino acids Type: general – SubjectFull: Cyanophycin Type: general – SubjectFull: Ligases Type: general – SubjectFull: Enzymes Type: general Titles: – TitleFull: Investigations on three genes in Ralstonia eutropha H16 encoding putative cyanophycin metabolizing enzymes. Type: main BibRelationships: HasContributorRelationships: – PersonEntity: Name: NameFull: Adames, Katja – PersonEntity: Name: NameFull: Euting, Karina – PersonEntity: Name: NameFull: Bröker, Anna – PersonEntity: Name: NameFull: Steinbüchel, Alexander IsPartOfRelationships: – BibEntity: Dates: – D: 15 M: 04 Text: Apr2013 Type: published Y: 2013 Identifiers: – Type: issn-print Value: 01757598 Numbering: – Type: volume Value: 97 – Type: issue Value: 8 Titles: – TitleFull: Applied Microbiology & Biotechnology Type: main |
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