A single amino acid substitution in aromatic hydroxylase (HpaB) of Escherichia coli alters substrate specificity of the structural isomers of hydroxyphenylacetate
Abstract Background A broad range of aromatic compounds can be degraded by enteric bacteria, and hydroxyphenylacetic acid (HPA) degrading bacteria are the most widespread. Majority of Escherichia coli strains can use both the structural isomers of HPA, 3HPA and 4HPA, as the sole carbon source, which...
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BMC
2020-05-01
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| Series: | BMC Microbiology |
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| Online Access: | http://link.springer.com/article/10.1186/s12866-020-01798-4 |
| _version_ | 1818445088296009728 |
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| author | Hanseol Kim Sinyeon Kim Dohyeon Kim Sung Ho Yoon |
| author_facet | Hanseol Kim Sinyeon Kim Dohyeon Kim Sung Ho Yoon |
| author_sort | Hanseol Kim |
| collection | DOAJ |
| description | Abstract Background A broad range of aromatic compounds can be degraded by enteric bacteria, and hydroxyphenylacetic acid (HPA) degrading bacteria are the most widespread. Majority of Escherichia coli strains can use both the structural isomers of HPA, 3HPA and 4HPA, as the sole carbon source, which are catabolized by the same pathway whose associated enzymes are encoded by hpa gene cluster. Previously, we observed that E. coli B REL606 grew only on 4HPA, while E. coli B BL21(DE3) grew on 3HPA as well as 4HPA. Results In this study, we report that a single amino acid in 4-hydroxyphenylacetate 3-hydroxylase (HpaB) of E. coli determines the substrate specificity of HPA isomers. Alignment of protein sequences encoded in hpa gene clusters of BL21(DE3) and REL606 showed that there was a difference of only one amino acid (position 379 in HpaB) between the two, viz., Arg in BL21(DE3) and Cys in REL606. REL606 cells expressing HpaB having Arg379 could grow on 3HPA, whereas those expressing HpaB with Gly379 or Ser379 could not. Structural analysis suggested that the amino acid residue at position 379 of HpaB is located not in the active site, but in the vicinity of the 4HPA binding site, and that it plays an important role in mediating the entrance and stable binding of substrates to the active site. Conclusions The arginine residue at position 379 of HpaB is critical for 3HPA recognition. Information regarding the effect of amino acid residues on the substrate specificity of structural isomers can facilitate in designing hydoxylases with high catalytic efficiency and versatility. |
| first_indexed | 2024-12-14T19:26:16Z |
| format | Article |
| id | doaj.art-2f8140aed4f34081ac21d3d5f5d25ded |
| institution | Directory Open Access Journal |
| issn | 1471-2180 |
| language | English |
| last_indexed | 2024-12-14T19:26:16Z |
| publishDate | 2020-05-01 |
| publisher | BMC |
| record_format | Article |
| series | BMC Microbiology |
| spelling | doaj.art-2f8140aed4f34081ac21d3d5f5d25ded2022-12-21T22:50:11ZengBMCBMC Microbiology1471-21802020-05-012011910.1186/s12866-020-01798-4A single amino acid substitution in aromatic hydroxylase (HpaB) of Escherichia coli alters substrate specificity of the structural isomers of hydroxyphenylacetateHanseol Kim0Sinyeon Kim1Dohyeon Kim2Sung Ho Yoon3Department of Bioscience and Biotechnology, Konkuk UniversityDepartment of Bioscience and Biotechnology, Konkuk UniversityDepartment of Bioscience and Biotechnology, Konkuk UniversityDepartment of Bioscience and Biotechnology, Konkuk UniversityAbstract Background A broad range of aromatic compounds can be degraded by enteric bacteria, and hydroxyphenylacetic acid (HPA) degrading bacteria are the most widespread. Majority of Escherichia coli strains can use both the structural isomers of HPA, 3HPA and 4HPA, as the sole carbon source, which are catabolized by the same pathway whose associated enzymes are encoded by hpa gene cluster. Previously, we observed that E. coli B REL606 grew only on 4HPA, while E. coli B BL21(DE3) grew on 3HPA as well as 4HPA. Results In this study, we report that a single amino acid in 4-hydroxyphenylacetate 3-hydroxylase (HpaB) of E. coli determines the substrate specificity of HPA isomers. Alignment of protein sequences encoded in hpa gene clusters of BL21(DE3) and REL606 showed that there was a difference of only one amino acid (position 379 in HpaB) between the two, viz., Arg in BL21(DE3) and Cys in REL606. REL606 cells expressing HpaB having Arg379 could grow on 3HPA, whereas those expressing HpaB with Gly379 or Ser379 could not. Structural analysis suggested that the amino acid residue at position 379 of HpaB is located not in the active site, but in the vicinity of the 4HPA binding site, and that it plays an important role in mediating the entrance and stable binding of substrates to the active site. Conclusions The arginine residue at position 379 of HpaB is critical for 3HPA recognition. Information regarding the effect of amino acid residues on the substrate specificity of structural isomers can facilitate in designing hydoxylases with high catalytic efficiency and versatility.http://link.springer.com/article/10.1186/s12866-020-01798-4Hydroxyphenylacetic acid4-hydroxyphenylacetate 3-hydroxylaseStructural isomerSubstrate specificity |
| spellingShingle | Hanseol Kim Sinyeon Kim Dohyeon Kim Sung Ho Yoon A single amino acid substitution in aromatic hydroxylase (HpaB) of Escherichia coli alters substrate specificity of the structural isomers of hydroxyphenylacetate BMC Microbiology Hydroxyphenylacetic acid 4-hydroxyphenylacetate 3-hydroxylase Structural isomer Substrate specificity |
| title | A single amino acid substitution in aromatic hydroxylase (HpaB) of Escherichia coli alters substrate specificity of the structural isomers of hydroxyphenylacetate |
| title_full | A single amino acid substitution in aromatic hydroxylase (HpaB) of Escherichia coli alters substrate specificity of the structural isomers of hydroxyphenylacetate |
| title_fullStr | A single amino acid substitution in aromatic hydroxylase (HpaB) of Escherichia coli alters substrate specificity of the structural isomers of hydroxyphenylacetate |
| title_full_unstemmed | A single amino acid substitution in aromatic hydroxylase (HpaB) of Escherichia coli alters substrate specificity of the structural isomers of hydroxyphenylacetate |
| title_short | A single amino acid substitution in aromatic hydroxylase (HpaB) of Escherichia coli alters substrate specificity of the structural isomers of hydroxyphenylacetate |
| title_sort | single amino acid substitution in aromatic hydroxylase hpab of escherichia coli alters substrate specificity of the structural isomers of hydroxyphenylacetate |
| topic | Hydroxyphenylacetic acid 4-hydroxyphenylacetate 3-hydroxylase Structural isomer Substrate specificity |
| url | http://link.springer.com/article/10.1186/s12866-020-01798-4 |
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