Publication Details |
| Category | Text Publication |
| Reference Category | Journals |
| DOI | 10.1128/AEM.00716-15 |
| Title (Primary) | Thermophilic coenzyme B12-dependent acyl-coenzyme A (CoA) mutase from Kyrpidia tusciae DSM 2912 preferentially catalyzes isomerization of (R)-3-hydroxybutyryl- and 2-hydroxyisobutyryl-CoA |
| Author | Weichler, M.-T.; Kurteva-Yaneva, N.; Przybylski, D.; Schuster, J.; Müller, R.H.; Harms, H.; Rohwerder, T. |
| Source Titel | Applied and Environmental Microbiology |
| Year | 2015 |
| Department | UMB |
| Volume | 81 |
| Issue | 14 |
| Page From | 4564 |
| Page To | 4572 |
| Language | englisch |
| Supplements | https://aem.asm.org/highwire/filestream/59629/field_highwire_adjunct_files/0/zam999116365so1.pdf |
| UFZ wide themes | RU4; |
| Abstract | The recent discovery of a coenzyme B12-dependent acyl-coenzyme A (CoA) mutase isomerizing 3-hydroxybutyryl- and 2-hydroxyisobutyryl-CoA in the mesophilic bacterium Aquincola tertiaricarbonis L108 (Yaneva et al., J. Biol. Chem., 287:15502-15511, 2012) could pave the way for a complete biosynthesis route to the building block chemical 2-hydroxyisobutyric acid from renewable carbon. However, the enzyme catalyzes only the conversion of the stereoisomer (S)-3-hydroxybutyryl-CoA at reasonable rates which seriously hampers an efficient combination of mutase and well-established bacterial poly-(R)-3-hydroxybutyrate (PHB) overflow metabolism. Here, we characterize a new 2-hydroxyisobutyryl-CoA mutase found in the thermophilic knallgas bacterium Kyrpidia tusciae DSM 2912. Reconstituted mutase subunits revealed highest activity at 55°C. Surprisingly, already at 30°C isomerization of (R)-3-hydroxybutyryl-CoA was about 7,000 times more efficient than with the mutase from strain L108. The most striking structural difference between the two mutases, likely determining stereospecificity, is a substitution of active site residue Asp found in strain L108 at position 117 with Val in the enzyme from strain DSM 2912, resulting in a reversed polarity at this binding site. Overall sequence comparison indicates that both enzymes descended from different prokaryotic thermophilic methylmalonyl-CoA mutases. Concomitant expression of PHB enzymes delivering (R)-3-hydroxybutyryl-CoA (beta-ketothiolase PhaA and acetoacetyl-CoA reductase PhaB from Cupriavidus necator) with the new mutase in E. coli JM109 and BL21 strains incubated on gluconic acid at 37°C led to the production of 2-hydroxyisobutyric acid at maximal titers of 0.7 mM. Measures to improve production in E. coli, such as co-expression of the chaperone MeaH and repression of thioesterase II, are discussed. |
| Persistent UFZ Identifier | https://www.ufz.de/index.php?en=20939&ufzPublicationIdentifier=16093 |
| Weichler, M.-T., Kurteva-Yaneva, N., Przybylski, D., Schuster, J., Müller, R.H., Harms, H., Rohwerder, T. (2015): Thermophilic coenzyme B12-dependent acyl-coenzyme A (CoA) mutase from Kyrpidia tusciae DSM 2912 preferentially catalyzes isomerization of (R)-3-hydroxybutyryl- and 2-hydroxyisobutyryl-CoA Appl. Environ. Microb. 81 (14), 4564 - 4572 10.1128/AEM.00716-15 |
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