Topic
CoA-transferase activity
About: CoA-transferase activity is a research topic. Over the lifetime, 7 publications have been published within this topic receiving 121 citations.
Papers
TL;DR: The dehydrogenase and the CoA transferase fit well into the proposed pathway of leucine reduction to isocaproate, and the conserved D171 was identified as an essential catalytic residue probably involved in the formation of a mixed anhydride with the acyl group of the thioester substrate.
Abstract: The strictly anaerobic pathogenic bacterium Clostridium difficile occurs in the human gut and is able to thrive from fermentation of leucine. Thereby the amino acid is both oxidized to isovalerate plus CO2 and reduced to isocaproate. In the reductive branch of this pathway, the dehydration of (R)-2-hydroxyisocaproyl-coenzyme A (CoA) to (E)-2-isocaprenoyl-CoA is probably catalyzed via radical intermediates. The dehydratase requires activation by an ATP-dependent one-electron transfer (J. Kim, D. Darley, and W. Buckel, FEBS J. 272:550-561, 2005). Prior to the dehydration, a dehydrogenase and a CoA transferase are supposed to be involved in the formation of (R)-2-hydroxyisocaproyl-CoA. Deduced amino acid sequences of ldhA and hadA from the genome of C. difficile showed high identities to d-lactate dehydrogenase and family III CoA transferase, respectively. Both putative genes encoding the dehydrogenase and CoA transferase were cloned and overexpressed in Escherichia coli; the recombinant Strep tag II fusion proteins were purified to homogeneity and characterized. The substrate specificity of the monomeric LdhA (36.5 kDa) indicated that 2-oxoisocaproate (Km = 68 μM, kcat = 31 s−1) and NADH were the native substrates. For the reverse reaction, the enzyme accepted (R)- but not (S)-2-hydroxyisocaproate and therefore was named (R)-2-hydroxyisocaproate dehydrogenase. HadA showed CoA transferase activity with (R)-2-hydroxyisocaproyl-CoA as a donor and isocaproate or (E)-2-isocaprenoate as an acceptor. By site-directed mutagenesis, the conserved D171 was identified as an essential catalytic residue probably involved in the formation of a mixed anhydride with the acyl group of the thioester substrate. However, neither hydroxylamine nor sodium borohydride, both of which are inactivators of the CoA transferase, modified this residue. The dehydrogenase and the CoA transferase fit well into the proposed pathway of leucine reduction to isocaproate.
68 citations
TL;DR: Growth of the bamY mutant with benzoate can be rationalized by the identification of a previously unknown succinyl-CoA:benzoate CoA transferase activity, which represents an additional, energetically less demanding mode of benzosate activation.
Abstract: Geobacter metallireducens is a Fe(III)-respiring deltaproteobacterium and serves as a model organism for aromatic compound-degrading, obligately anaerobic bacteria. In this study, a genetic system was established for G. metallireducens using nitrate as an alternative electron acceptor. Surprisingly, disruption of the benzoate-induced bamY gene, encoding a benzoate coenzyme A (CoA) ligase, reproducibly showed an increased biomass yield in comparison to the wild type during growth with benzoate but not during growth with acetate. Complementation of bamY in trans converted the biomass yield back to the wild-type level. Growth of the bamY mutant with benzoate can be rationalized by the identification of a previously unknown succinyl-CoA:benzoate CoA transferase activity; it represents an additional, energetically less demanding mode of benzoate activation. The activity was highly enriched from extracts of cells grown on benzoate, yielding a 50-kDa protein band; mass spectrometric analysis identified the corresponding benzoate-induced gene annotated as a CoA transferase. It was heterologously expressed in Escherichia coli and characterized as a specific succinyl-CoA:benzoate CoA transferase. The newly identified enzyme in conjunction with a benzoate-induced succinyl-CoA synthetase links the tricarboxylic acid cycle to the upper benzoyl-CoA degradation pathway during growth on aromatic growth substrates.
27 citations
Journal Article•
TL;DR: Results clearly indicate that the faster-growing hepatomas have adequate capacity to utilize ketone bodies in bioenergetic or biosynthetic activities.
Abstract: The presence of succinyl-coenzyme A:acetoacetate CoA transferase (CoA transferase) (EC 2.8.3.5), an initiator of ketone body utilization in nonhepatic tissue, was examined in liver from normal, partly hepatectomized, neonatal, and tumor-bearing rats, as well as in a series of transplantable rat hepatomas ranging widely in growth rate. While levels of CoA transferase are extremely low in normal, host, and regenerating liver, considerable amounts of activity are detectable in neonatal liver and in the hepatomas. In fact, the content of CoA transferase in the series of Morris hepatomas increases progressively with increase in tumor-growth rate. The fastest-growing tumor studied (7288Ctc) contains about the same amount of CoA transferase activity as rat skeletal muscle (i.e., an activity of about 0.1 mumole of acetoacetate used per min per g tissue). These results clearly indicate that the faster-growing hepatomas have adequate capacity to utilize ketone bodies in bioenergetic or biosynthetic activities. Furthermore, the enzymes from normal and hepatoma 7288Ctc tissues are quite similar with respect to (a) size of about 10(5) daltons, (b) reaction mechanism requiring formation of an enzyme:CoA intermediate (from ping-pong kinetic data), and (c) various kinetic parameters (such as Michaelis constants, product competitive inhibition constants, and acetoacetate substrate inhibition). The enzymes from rat skeletal muscle and Morris hepatoma 7288Ctc have the same isoelectric point (7.6), which differs from that for the rat heart enzyme (6.8).
20 citations
Journal Article•
TL;DR: The expression of CoA transferase in hepatoma cells may play a role in energy production because of the varying amounts of the enzyme synthesized in the cells as monitored by L-[35S]methionine labeling followed by immunoprecipitation.
Abstract: The regulation of succinyl-CoA:acetoacetyl-CoA transferase (CoA transferase) has been studied in 8 rat hepatoma cell lines. Compared with normal rat hepatocytes, which have almost nondetectable activity of the enzyme, the hepatoma cell lines have a wide range of expression of CoA transferase activity, from as low as 45 nmol/min/mg to as high as 960 nmol/min/mg. Western blotting showed that the different levels of CoA transferase activity were due to differing amounts of the enzyme in the cells. This was further attributed to the varying amounts of the enzyme synthesized in the cells as monitored by L-[35S]methionine labeling followed by immunoprecipitation. To study further the differential expression of CoA transferase in the hepatoma cell lines, the relative quantity of functional CoA-transferase mRNA in the cells was measured by in vitro translation. The results showed that the levels of functional CoA transferase mRNA detected were consistent with the differences in the enzyme activity in the cells. Since CoA transferase is the key enzyme responsible for the utilization of ketone bodies as an alternative energy source, the expression of CoA transferase in hepatoma cells may play a role in energy production.
11 citations
TL;DR: The competitive inhibition of the product succinate with respect to both reactants, 3-hydroxy-3-methylglutarate and succinyl-CoA, as well as the Haldane relationships are consistent with this conclusion.
7 citations
Related Topics (5)
Performance Metrics
| Year | Papers |
|---|---|
| 2014 | 1 |
| 2012 | 1 |
| 2006 | 1 |
| 1990 | 1 |
| 1989 | 1 |
| 1984 | 1 |