Actinobacterial Degradation of 2-Hydroxyisobutyric Acid Proceeds via Acetone and Formyl-CoA by Employing a Thiamine-Dependent Lyase Reaction Rohwerder, Thore Rohde, Maria-Teresa Jehmlich, Nico Purswani, Jessica Degradation pathway Isobutene Tert-butyl alcohol Fuel oxygenate Mycolicibacterium 2-hydroxyacyl-CoA lyase Acyloin condensation We would like to thank C. Dilßner and M. Neytschev (UFZ) for excellent technical assistance with CoA thioester synthesis, strain cultivation and HPLC analyses. In addition, we thank Birgit Würz (UFZ) for invaluable analytical advice and help with GC mass spectrometry. We are also indebted to L. von Wintzingerode, A. Grunwald, and J. Grabengießer (UFZ) for assistance in the cultivation and enzyme assay experiments. Many thanks to K. Eismann (UFZ) as well, for help with the proteome analysis and fruitful discussions regarding different protein extraction methods. The Supplementary Material for this article can be found online at: https://www.frontiersin.org/articles/10.3389/fmicb.2020.00691/full#supplementary-material The tertiary branched short-chain 2-hydroxyisobutyric acid (2-HIBA) has been associated with several metabolic diseases and lysine 2-hydroxyisobutyrylation seems to be a common eukaryotic as well as prokaryotic post-translational modification in proteins. In contrast, the underlying 2-HIBA metabolism has thus far only been detected in a few microorganisms, such as the betaproteobacterium Aquincola tertiaricarbonis L108 and the Bacillus group bacterium Kyrpidia tusciae DSM 2912. In these strains, 2-HIBA can be specifically activated to the corresponding CoA thioester by the 2-HIBA-CoA ligase (HCL) and is then isomerized to 3-hydroxybutyryl-CoA in a reversible and B12-dependent mutase reaction. Here, we demonstrate that the actinobacterial strain Actinomycetospora chiangmaiensis DSM 45062 degrades 2-HIBA and also its precursor 2-methylpropane-1,2-diol via acetone and formic acid by employing a thiamine pyrophosphate-dependent lyase. The corresponding gene is located directly upstream of hcl, which has previously been found only in operonic association with the 2-hydroxyisobutyryl-CoA mutase genes in other bacteria. Heterologous expression of the lyase gene from DSM 45062 in E. coli established a 2-hydroxyisobutyryl-CoA lyase activity in the latter. In line with this, analysis of the DSM 45062 proteome reveals a strong induction of the lyase-HCL gene cluster on 2-HIBA. Acetone is likely degraded via hydroxylation to acetol catalyzed by a MimABCD-related binuclear iron monooxygenase and formic acid appears to be oxidized to CO2 by selenium-dependent dehydrogenases. The presence of the lyase-HCL gene cluster in isoprene-degrading Rhodococcus strains and Pseudonocardia associated with tropical leafcutter ant species points to a role in degradation of biogenic short-chain ketones and highly branched organic compounds. 2020-06-01T11:46:33Z 2020-06-01T11:46:33Z 2020-04-15 info:eu-repo/semantics/article Rohwerder T, Rohde M-T, Jehmlich N and Purswani J (2020) Actinobacterial Degradation of 2-Hydroxyisobutyric Acid Proceeds via Acetone and Formyl-CoA by Employing a Thiamine-Dependent Lyase Reaction. Front. Microbiol. 11:691. [doi: 10.3389/fmicb.2020.00691] http://hdl.handle.net/10481/62312 10.3389/fmicb.2020.00691 eng EC/FP7/624857 http://creativecommons.org/licenses/by/3.0/es/ info:eu-repo/semantics/openAccess Atribución 3.0 España Frontiers Media