Ethylmalonyl CoA route

The ethylmalonyl-CoA pathway is a metabolic pathway in some bacteria that is used to assimilate acetate . Here, three molecules of acetate, one molecule of carbon dioxide and one molecule of hydrogen carbonate are built up to form the citric acid cyclite medium L - malate and succinyl-CoA . The ethylmalonyl-CoA route is an alternative to the glyoxylate or methylaspartate cycle .

Occurrence

The metabolic pathway was found in some bacteria that lack an important enzyme, isocitrate lyase ( EC  4.1.3.1 ), to operate the glyoxylate cycle, for example in Rhodobacter sphaeroides , Methylobacterium exotrquens and Streptomyces coelicolor . These are representatives of non-sulfur purple bacteria and Alphaproteobacteria . Sequence comparisons of key enzymes of the ethylmalonyl-CoA pathway indicate that at least 38 types of bacteria can operate this metabolic pathway.

biochemistry

The ethylmalonyl-CoA pathway can be divided into three sections, whereby the reactions of the first and the third are not only typical for this metabolic pathway.

Formation of crotonyl-CoA from two molecules of acetyl-CoA

The path starts with the condensation of two molecules of acetyl-CoA ( 1 , see figure) to acetoacetyl-CoA ( 2 ), which catalyzes a β-ketothiolase . Acetoacetyl-CoA is then reduced to ( R ) -3-hydroxybutyryl-CoA ( 3 ) with NADPH consumption by an acetoacetyl-CoA reductase . The subsequent elimination of water can either take place directly or after ( R ) -3-hydroxybutyryl-CoA has first been converted to the S isomer . In both cases, crotonyl-CoA is formed ( 4 ).

This reaction sequence corresponds to the first steps in the formation of the bacterial storage substance polyhydroxybutyrate .

Conversion of crotonyl-CoA to mesaconyl-CoA

Crotonyl-CoA is then reductively by a crotonyl-CoA carboxylase / reductase (Ccr) carboxylated , wherein NADPH is consumed. This creates (2 S ) -ethylmalonyl-CoA (5). This is converted to the R-isomer ( 6 ) by a bifunctional epimerase (ethylmalonyl-CoA / methylmalonyl-CoA- epimerase ), which is converted to (2nd ) by a vitamin B12 -dependent (2 R ) -ethylmalonyl-CoA mutase (Ecm) S ) -Methylsuccinyl-CoA ( 7 ) reacts. A (2 S ) -Methylsuccinyl-CoA dehydrogenase (Mcd) finally oxidizes this product to mesaconyl- (C1) -CoA ( 8 ), the electron acceptor is not yet known.

The conversion of crotonyl-CoA to mesaconyl-CoA is unique and typical for this metabolic pathway.

Conversion of mesaconyl-CoA and acetyl-CoA to L- malate and succinyl-CoA

Water is then added to mesaconyl-CoA, which catalyzes a mesaconyl-CoA hydratase. (2 R , 3 S ) -β-methylmalyl-CoA ( 9 ) is formed. This is then cleaved into glyoxylate ( 10 ) and propionyl-CoA ( 13 ) by a β-methylmalyl-CoA / (3 S ) -malyl-CoA lyase. The type of reaction here is a Claisen condensation . Propionyl-CoA is carboxylated with consumption of ATP , which catalyzes a propionyl-CoA carboxylase . The resulting (2 S ) -methylmalonyl-CoA ( 14 ) is converted into its R-isomer ( 15 ) by the bifunctional epimerase (ethylmalonyl-CoA / methylmalonyl-CoA-epimerase) already described above and then by a mutase to succinyl -CoA ( 16 ) reshaped.

Another molecule of acetyl-CoA condenses on glyoxylate, so that (3 S ) -malyl-CoA ( 11 ) is formed. This reaction is also catalyzed by β-methylmalyl-CoA / (3 S ) -malyl-CoA lyase. In order for the reaction to proceed in the direction of (3 S ) malate (= L malate ) ( 12 ), the thioester must still be irreversibly cleaved. This is finally done by a (3 S ) -malyl-CoA thioesterase. This enzyme was recently detected in R. sphaeroides . There the comparison with the precursor enzyme shows that it has 33% sequence homology, although a completely different type of reaction is present. In addition, the enzyme is very substrate-specific.

Some of these reactions are also not limited to the ethylmalonyl-CoA route. The carboxylation of propionyl-CoA is carried out by many organisms in the course of the methylmalonate route . The catalyzed reactions of β-methylmalyl-CoA / (3 S ) -malyl-CoA lyase in the 3-hydroxypropionate cycle are represented in reverse order : there, malyl-CoA is split into acetyl-CoA and glyoxylate and the latter condenses with propionyl-CoA.

The ethylmalonyl-CoA pathway at a glance. The unique metabolites and enzymes typical of this metabolic pathway are highlighted in blue. Please see text for details.

Balance sheet

In the ethylmalonyl-CoA pathway, three molecules of acetyl-CoA, one molecule of carbon dioxide and one molecule of hydrogen carbonate are built up into one molecule each of succinyl-CoA and L-malate:

${\displaystyle \mathrm {3\ C_{2}+2\ C_{1}\longrightarrow 2\ C_{4}} }$

If you take into account other factors, such as the reduction equivalents, the total balance is:

${\displaystyle \mathrm {3\ Acetyl{\text{-}}CoA+CO_{2}+HCO_{3}^{-}+2\ NADPH+2\ H^{+}+ATP+H_{2}O\longrightarrow } }$

${\displaystyle \mathrm {Succinyl{\text{-}}CoA+Malat+2\ HS{\text{-}}CoA+2\ NADP^{+}+2\ [H]+ADP+P_{i}} }$

The carbon dioxide does not necessarily have to come from the environment. R. sphaeroides , for example, also grows with acetate as the only carbon source. The CO ₂ from the citric acid cycle is probably used there , which is released by the metabolism of acetyl-CoA.

Biological importance

In many organisms, acetate is assimilated exclusively via the glyoxylate cycle and converted into metabolic intermediates such as succinyl-CoA or malate. Some microorganisms are known to be unable to operate the glyoxylate cycle because they lack an important key enzyme, isocitrate lyase. Nevertheless, it has been observed that these bacteria can grow on acetate and therefore have a different assimilation strategy: the ethylmalonyl-CoA pathway.

The ethylmalonyl-CoA pathway allows those microorganisms to assimilate acetate to build L- malate and succinyl-CoA. From this, other metabolites such as amino acids can be used to build cell material. Since these are intermediates in the citric acid cycle, the ethylmalonyl-CoA pathway is an anaplerotic reaction .

The ethylmalonyl-CoA pathway can also be linked to other metabolic pathways. In the case of methanotrophs of type II, which do not have any isocitrate lyase, acetyl-CoA is produced from C ₁ components such as formaldehyde or methanol in the course of the serine pathway . Two generated molecules of acetyl-CoA could flow into the ethylmalonyl-CoA pathway and be built up to succinyl-CoA and glyoxylate. Glyoxylate then gets back into the serine pathway and is built up into PEP , which can be withdrawn to build up carbohydrates.

${\displaystyle \mathrm {3\ C_{1}+3\ CO_{2}+HCO_{3}^{-}\longrightarrow PEP+Succinyl{\text{-}}CoA} }$

Other variations are also conceivable, for example a cycle in which 3-phosphoglycerate is produced from the above-mentioned C ₁ units.

The ethylmalonyl-CoA route is not viewed as an exception, but as an alternative to the glyoxylate cycle, as at least 57 types of bacteria have the enzymes required for this route. There are also at least nine types of bacteria that are believed to be able to use either route of acetate assimilation. For example, Paracoccus versutus , an optional denitrifier , probably operates the ethylmalonyl-CoA pathway under aerobic conditions, but the glyoxylate cycle under anaerobic conditions.

While in the glyoxylate cycle one enzyme, malate synthase, catalyzes the conversion of glyloxylate with acetyl-CoA to L -malate, two enzymes are required in the ethylmalonyl-CoA pathway.

However, some halobacteria , such as Haloarcula marismortui , lack both the key enzymes for the ethylmalonyl-CoA pathway and the isocitrate lyase for the glyoxylate cycle. It has been shown that a third route to acetate assimilation exists, the so-called methylaspartate cycle . A total of nine reaction steps are required to convert isocitrate to succinate. This creates the eponymous methyl aspartate, an unusual, non-proteinogenic amino acid .

Individual evidence

1. ↑ Erb, TJ. et al . (2010): The apparent malate synthase activity of Rhodobacter sphaeroides is due to two paralogous enzymes, (3S) -Malyl-coenzyme A (CoA) / {beta} -methylmalyl-CoA lyase and (3S) - Malyl-CoA thioesterase . In: J Bacteriol. 192 (5); 1249-1258; PMID 20047909 ; doi : 10.1128 / JB.01267-09
2. ↑ ^{a b} Erb, TJ. et al . (2007): Synthesis of C5-dicarboxylic acids from C2-units involving crotonyl-CoA carboxylase / reductase: the ethylmalonyl-CoA pathway . In: Proc Natl Acad Sci USA 104 (25); 10631-10636; PMID 17548827 ; PMC 1965564 (free full text)
3. ↑ Peyraud, R. et al . (2009): Demonstration of the ethylmalonyl-CoA pathway by using 13C metabolomics . In: Proc Natl Acad Sci USA . 106 (12); 4846-4851; PMID 19261854 ; PMC 2660752 (free full text)
4. ↑ Khomyakova, M., Bükmez, Ö., Thomas, LK., Erb, TJ. and Berg, IA. (2011): A methylaspartate cycle in haloarchaea . In: Science 331 (6015); 334-337; PMID 21252347 ; doi : 10.1126 / science.1196544

literature

• Erb, TJ., Fuchs, G. and Alber, BE. (2009): (2S) -Methylsuccinyl-CoA dehydrogenase closes the ethylmalonyl-CoA pathway for acetyl-CoA assimilation . In: Mol Microbiol . 73 (6); 992-1008; PMID 19703103 ; doi : 10.1111 / j.1365-2958.2009.06837.x
• Schink, B. (2009): An alternative to the glyoxylate shunt . In: Mol Microbiol . 73 (6); 975-977; PMID 19682245 ; doi : 10.1111 / j.1365-2958.2009.06835.x

Web links

• Alternative acetate assimilation routes AG Fuchs, University of Freiburg