Acrylyl-Coenzyme A Reductase, an Enzyme Involved in the Assimilation of 3-Hydroxypropionate by Rhodobacter sphaeroides
| Autor: | Birgit E. Alber, Marie Asao |
|---|---|
| Rok vydání: | 2013 |
| Předmět: |
Coenzyme A
Ruegeria Dehydrogenase Rhodobacter sphaeroides Reductase medicine.disease_cause Microbiology Gene Expression Regulation Enzymologic chemistry.chemical_compound Bacterial Proteins medicine Histidine Lactic Acid Molecular Biology Escherichia coli chemistry.chemical_classification biology Gene Expression Regulation Bacterial Articles biology.organism_classification Enzyme assay Enzyme chemistry Biochemistry biology.protein Oxidoreductases |
| Zdroj: | Journal of Bacteriology. 195:4716-4725 |
| ISSN: | 1098-5530 0021-9193 |
| DOI: | 10.1128/jb.00685-13 |
| Popis: | The anoxygenic phototroph Rhodobacter sphaeroides uses 3-hydroxypropionate as a sole carbon source for growth. Previously, we showed that the gene (RSP_1434) known as acuI , which encodes a protein of the medium-chain dehydrogenase/reductase (MDR) superfamily, was involved in 3-hydroxypropionate assimilation via the reductive conversion to propionyl-coenzyme A (CoA). Based on these results, we speculated that acuI encoded acrylyl-CoA reductase. In this work, we characterize the in vitro enzyme activity of purified, recombinant AcuI using a coupled spectrophotometric assay. AcuI from R. sphaeroides catalyzes the NADPH-dependent acrylyl-CoA reduction to produce propionyl-CoA. Two other members of the MDR012 family within the MDR superfamily, the products of SPO_1914 from Ruegeria pomeroyi and yhdH from Escherichia coli , were shown to also be part of this new class of NADPH-dependent acrylyl-CoA reductases. The activities of the three enzymes were characterized by an extremely low K m for acrylyl-CoA (−1 . These homodimeric enzymes were highly specific for NADPH ( K m = 18 to 33 μM), with catalytic efficiencies of more than 10-fold higher for NADPH than for NADH. The introduction of codon-optimized SPO_1914 or yhdH into a Δ acuI :: kan mutant of R. sphaeroides on a plasmid complemented 3-hydroxypropionate-dependent growth. However, in their native hosts, SPO_1914 and yhdH are believed to function in the metabolism of substrates other than 3-hydroxypropionate, where acrylyl-CoA is an intermediate. Complementation of the Δ acuI :: kan mutant phenotype by crotonyl-CoA carboxylase/reductase from R. sphaeroides was attributed to the fact that the enzyme also uses acrylyl-CoA as a substrate. |
| Databáze: | OpenAIRE |
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