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Published in final edited form as: Clin Chim Acta. 2025 Apr 17;574:120320. doi: 10.1016/j.cca.2025.120320

Metabolic Origin and Significance of 3-Methylglutaryl CoA

Elizabeth A Jennings 1, Zane H Abi-Rached 1, Robert O Ryan 1
PMCID: PMC12094887  NIHMSID: NIHMS2075943  PMID: 40252717

Abstract

3-Methylglutaryl (3MG) CoA is not part of any biochemical pathway, yet its byproducts, 3MG carnitine and 3MG acid, are disease biomarkers. Both compounds are excreted in HMG CoA lyase deficiency, while 3MG aciduria occurs in inborn errors of metabolism (IEM) associated with compromised mitochondrial energy metabolism. In one such disorder (i.e., TMEM70 deficiency), 3MG carnitine is also present. Moreover, in a number of chronic and acute maladies, elevated levels of 3MG carnitine are present. The precursor of 3MG CoA is trans-3-methylglutaconyl (3MGC) CoA. When trans-3MGC CoA levels rise, a portion of this metabolite pool is reduced to 3MG CoA, potentially via a side reaction involving glutaryl CoA dehydrogenase (GCDH), which normally catalyzes the oxidative decarboxylation of glutaryl CoA to crotonyl CoA and CO2. This reaction occurs via a two-step process wherein glutaryl CoA is initially oxidized to glutaconyl CoA, coupled to reduction of the enzyme’s FAD prosthetic group. Enzyme-bound glutaconyl CoA is then decarboxylated to the reaction product, crotonyl CoA. Before GCDH can accept another glutaryl CoA the flavin prosthetic group must be oxidized to FAD by donating electrons to electron transferring flavoprotein (ETF). However, genetic- or disease-induced defects in electron transport chain function can impede this reaction. We propose that trans-3MGC CoA is a substrate for reduced GCDH and, when glutaryl CoA and trans-3MGC CoA are present, GCDH is able to bypass ETF and cycle between oxidized and reduced states, producing crotonyl CoA and CO2 from glutaryl CoA, and 3MG CoA from trans-3MGC CoA.

Keywords: Glutaryl CoA dehydrogenase, mitochondria, biomarker, 3-methylglutaric acid, 3-methylglutaryl carnitine

1. Introduction

3-Methylglutaryl (3MG) CoA is an unusual metabolite. It is not an intermediate in any known biochemical pathway nor can it be metabolized to yield energy. The fact that it is generally not detected in normal tissue extracts raises questions about why it should be of interest at all. The answer to this question relates to the fact that 3MG CoA is likely produced in significant amounts in various pathological conditions. Once formed, 3MG CoA is converted to the organic acid, 3MG acid, or the dicarboxy acylcarnitine, 3MG carnitine, both of which are destined for excretion. As such, 3MG acid and 3MG carnitine serve as disease biomarkers [1,2]. Although it is evident that 3MG CoA can be synthesized, and the primary fate of its carbon skeleton is excretion as a waste product, questions exist about the mechanism whereby this metabolite is produced. By examining the occurrence of 3MG acid and/or 3MG carnitine in subjects with various disease conditions, insight with respect to the metabolic origin of 3MG CoA has been gained.

2. 3MG Acid and Leucine Catabolism Pathway Enzyme Deficiencies

One of the earliest reports of 3MG acid excretion came from Wysocki et al [3]. These authors were investigating the presence of abnormal organic acids in urine of a patient with a deficiency in 3-hydroxy-3-methylglutaryl (HMG) CoA lyase (EC 4.1.3.4). Some of these organic acids, including 3-methylglutaconic (3MGC) acid and HMG acid, are derived directly from intermediates in the leucine catabolism pathway, while others appear to be products of side reactions (Figure 1). For example, 3-hydroxyisovaleric (HIV) acid is considered to arise from enoyl CoA hydratase-mediated (EC 4.2.1.17) hydration of 3-methylcrotonyl CoA. Additionally, 3MG acid is generated by reduction of the double bond in trans-3MGC CoA. In cases of HMG CoA lyase deficiency, these latter organic acids are quantitatively less important than the first two organic acids.

Figure 1. Organic acids formed in response to IEMs affecting HMG CoA lyase.

Figure 1.

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The leucine catabolism pathway is depicted on the left with the red bar depicting an IEM in HMG CoA lyase. Under these conditions, leucine pathway intermediates are redirected to various organic acids which are destined for excretion. Enzyme names are in boxes; abbreviations: IVD = isovaleryl CoA dehydrogenase; 3MCCCase = 3-methylcrotonyl CoA carboxylase; AUH = trans-3MGC CoA hydratase; HMGCL = HMG CoA lyase; ECH = enoyl CoA hydratase. Unlabeled arrows indicate non-enzymatic chemical reactions while the arrow labeled “reductase” refers to a putative reductase enzyme.

Faull et al [4] offered two possible explanations for the increased excretion of 3MG acid in cases of HMG CoA lyase deficiency. They first proposed that the saturated leucine pathway intermediate, isovaleryl CoA, can serve as a substrate for the leucine pathway enzyme, 3-methylcrotonyl CoA carboxylase (EC 6.4.1.4). In this case, 3-methylcrotonyl CoA carboxylase would convert isovaleryl CoA into 3MG CoA while bypassing 3-methylcrotonyl CoA (see Figure 1). This explanation is unlikely, however, because increased excretion of 3MG acid does not occur in patients with isovaleric acidemia. In a second postulate, these authors proposed that trans-3MGC CoA can be reduced to 3MG CoA by a reversal of the reaction normally catalyzed by isovaleryl CoA dehydrogenase (EC 1.3.99.10) (i.e, conversion of isovaleryl CoA to 3-methylcrotonyl CoA) [5]. This hypothesis appears unlikely, however, given structural differences between these molecules (i.e., trans-3MGC CoA is a terminally carboxylated acyl CoA while isovaleryl CoA has an aliphatic side chain).

Subsequently, Duran et al [6] showed that 3MG acid is also excreted in inborn errors of metabolism (IEM) affecting another leucine catabolism enzyme, 3MGC CoA hydratase (AUH; EC 4.2.1.18). In this disorder, the block in leucine catabolism occurs one step earlier than that seen in HMG CoA lyase deficiency (see Figure 1). Consistent with this, in AUH deficiency, HMG acid excretion is not observed. In both HMG CoA lyase and AUH deficiencies, leucine administration induces increased organic acid excretion [7].

3. 3MG Aciduria in IEMs Affecting Electron Transport Chain Function

In addition to deficiencies in HMG CoA lyase and AUH, (termed primary 3MGC acidurias) 3MG acid excretion is also a phenotypic feature of several discrete IEMs associated with compromised mitochondrial energy metabolism [8]. In these disorders, referred to as secondary 3MGC aciduria, excretion of 3MGC acid is also a prominent feature, and this organic acid is always more abundant than 3MG acid. Unlike the IEMs that cause primary 3MGC aciduria, 3HIV acid is not observed in secondary 3MGC aciduria [9]. Authors describing specific cases of secondary 3MGC aciduria originally proposed that an “isoprene shunt” is involved in shuttling isoprene units from cytosol to mitochondria [10,11]. This explanation, however, fails to account for the fact that, in every case, the mutated gene affects mitochondrial function. Subsequently, Su and Ryan [12] proposed that, in secondary 3MGC aciduria, trans-3MGC CoA is synthesized in mitochondria de novo from acetyl CoA rather than arising from either leucine catabolism or cytosolic isoprene units. Whereas such a pathway is known to exist in microorganisms [13], it had not been previously reported in human intermediary metabolism. The “acetyl CoA diversion pathway” (Figure 2) occurs in tissues with a high demand for ATP, such as cardiac and skeletal muscle mitochondria [1,14]. Briefly, above a threshold level of acetyl CoA, the reversible enzyme, acetoacetyl CoA thiolase (T2; EC 2.3.1.9), functions in reverse, catalyzing condensation of two acetyl CoA to form acetoacetyl CoA. Following this, HMG CoA synthase 2 (EC 2.3.3.10) catalyzes condensation of acetoacetyl CoA and acetyl CoA, yielding HMG CoA. Subsequently, HMG CoA is dehydrated by the reversible leucine pathway enzyme, AUH, forming trans-3MGC CoA. Once formed, this metabolic intermediate is unable to proceed further up the leucine catabolism pathway because the next reaction, catalyzed by 3-methylcrotonyl CoA carboxylase, is irreversible. Thus, under physiological conditions wherein acetyl CoA accumulates, it can be diverted from TCA cycle entry toward trans-3MGC CoA formation.

Figure 2. The acetyl CoA diversion pathway in IEMs that affect mitochondrial energy metabolism.

Figure 2.

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This pathway is driven by an accumulation of acetyl CoA in mitochondria and proceeds with two condensation reactions and a dehydration, forming trans-3MGC CoA. This leucine catabolism pathway intermediate is labile and can isomerize to cis-3MGC CoA (left side), followed by a series of non-enzymatic chemical reactions that yield cis-3MGC acid. Alternatively, trans-3MGC CoA can be reduced to 3MG CoA (right side) which is also susceptible to non-enzymatic intramolecular cyclization and hydrolysis to produce 3MG acid. Abbreviations: T2 Thiolase = acetoacetyl CoA thiolase; AUH = trans-3MGC CoA hydratase.

The acetyl CoA diversion pathway is initiated when there is a decline in TCA cycle activity due to any one of several IEMs that manifest functional defects in electron transport chain (ETC) function [8]. These defects impact one or more of the following: mitochondrial membrane lipid content and composition, inner membrane protein/enzyme function, and/or assembly of ATP synthase (EC 7.1.2.2). Directly or indirectly, these mutations affect a) electron flux through the ETC, b) establishment of a proton gradient, or c) productive coupling of an established proton motive force to ADP phosphorylation. When any of these functions are impaired, ATP synthesis rates decline, and aerobic energy metabolism is adversely affected. Under these physiological conditions, NADH and FADH2, generated during oxidative metabolism of fuel molecules, are unable to efficiently donate electrons to the ETC. As NADH levels rise in the mitochondrial matrix, end product inhibition of enzymes that generate this reduced cofactor during oxidative metabolism leads to inhibition of key metabolic pathways, including the TCA cycle. When this occurs, some portion of the trans-3MGC CoA pool is reduced to 3MG CoA via an apparent side reaction. Interestingly, Wortmann et al [7] noted differences in the ratio of 3MGC acid to 3MG acid in primary 3MGC aciduria versus those secondary 3MGC aciduria. The trend observed revealed that the ratio is lower in secondary 3MGC aciduria. Thus, despite the fact that significantly lower amounts of organic acids are generated in secondary 3MGC aciduria, the relative proportion of 3MG acid is higher.

4. Non-Enzymatic Intramolecular Cyclization of trans-3MGC CoA and 3MG CoA

As trans-3MGC CoA and 3MG CoA are produced, either via the acetyl CoA diversion pathway or by IEMs in the leucine degradation pathway, these metabolites have different potential fates. For example, trans-3MGC CoA can 1) be reduced to 3MG CoA by an unknown mechanism or 2) spontaneously isomerize to cis-3MGC CoA and undergo intramolecular cyclization to form 3MGC anhydride and free CoA. The anhydride can then be hydrolyzed, forming 3MGC acid (Figure 2). In addition, cyclic 3MGC anhydride can react with lysine side chain amino groups to covalently 3MGCylate proteins (Figure 3) [15,2]. 3) Finally, if physiological conditions normalize, trans-3MGC CoA can be metabolized via the leucine degradation pathway to acetoacetate and acetyl CoA.

Figure 3. Acylation of protein lysine side chain amino groups by trans-3MGC CoA and 3MG CoA.

Figure 3.

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Following their conversion to the corresponding cyclic anhydride, both cis-3MGC anhydride and 3MG anhydride can covalently acylate proteins by reacting with the side chain amino groups of lysine residues. The acyl groups can also be removed through the action of the NAD+-specific deacylase, sirtuin 4 (SIRT4), yielding the organic acids, 3MGC acid and 3MG acid, as products that are destined for excretion. Note that SIRT4 requires NAD+ as a substrate which is converted to nicotinamide and 2′-O-cis-3MGC-ADP-ribose or 2′-O-3MG-ADP-ribose as products (not shown). Subsequently, the 2′-O-cis-3MGC-ADP-ribose or 2′-O-3MG-ADP-ribose are hydrolyzed to yield ADP-ribose and the corresponding organic acid products depicted.

As a structurally related, terminally carboxylated, short-chain acyl CoA, when 3MG CoA is formed, it can also undergo spontaneous intramolecular cyclization, forming 3MG anhydride and free CoA (Figure 2). Furthermore, just as with cis-3MGC anhydride, 3MG anhydride is reactive and can either be hydrolyzed directly to 3MG acid or react with protein lysine side chain amino groups, forming a covalent adduct (i.e., protein 3MGylation) [16]. Importantly, protein 3MGCylation and 3MGylation, as depicted in figure 3, can be reversed by the NAD+-dependent protein deacylase, sirtuin 4 (EC 2.3.1.286) [17]. In addition, 3MG CoA has a unique alternate fate which is of potential biomedical relevance: conversion to 3MG carnitine (see below).

5. Conversion of 3MG CoA to 3MG Carnitine

3MG carnitine was originally identified as a diagnostic biomarker in patients with a deficiency in HMG CoA lyase [18]. As described above, HMG CoA lyase is a component of the leucine catabolism pathway, as well as ketone body synthesis in liver and kidneys. When this mitochondrial enzyme is deficient, a characteristic set of organic acids appear in urine, including HMG acid, 3MGC acid, 3HIV acid, and 3MG acid [19]. Unlike HMG CoA or trans-3MGC CoA, however, 3MG CoA is known to react with carnitine, forming 3MG carnitine [2], as shown in Figure 4. This short-chain dicarboxy acylcarnitine then exits the mitochondrial matrix to the cytosol, and then to the bloodstream, where it is readily detectable by mass spectrometry. Ultimately, 3MG carnitine is destined for excretion in urine. Roe et al [18] originally reported that 3MG CoA “is believed to originate” from reduction of the double bond in trans-3MGC CoA. Interestingly, HMG carnitine or trans-3MGC carnitine are rarely detected despite the fact that, in HMG CoA lyase deficiency, large amounts of HMG acid and 3MGC acid are excreted. Thus, it appears that the carnitine acyltransferase enzyme responsible for 3MG carnitine production does not recognize HMG CoA or trans-3MGC CoA as substrates. The human genome contains several distinct genes that encode enzymes with carnitine acyltransferase activity, including, but not limited to, carnitine acetyltransferase (CrAT, EC 2.3.1.7), carnitine palmitoyltransferase (CPT)-I (EC 2.3.1.21), CPT-II, and carnitine octanoyltransferase (CrOT; EC 2.3.1.137) [20]. At present, however, it is not clear which of these is responsible for 3MG carnitine formation. CrAT is located in the mitochondrial matrix and typically accepts substrates with chain lengths of 2 to 10 carbons. However, when Violante et al [21] investigated the substrate specificity of this enzyme, terminally carboxylated acyl CoAs were found to be poor substrates. Since 3MG CoA was not assayed in this study, a firm conclusion cannot be made about its potential role in 3MG carnitine formation.

Figure 4. Carnitine acyltransferase catalyzed conversion of 3MG CoA to 3MG carnitine.

Figure 4.

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The reversible enzymatic reaction, catalyzed by a carnitine acyltransferase (CAT) converts 3MG CoA and free carnitine to 3MG carnitine and CoA. The directionality of this reaction is dependent upon the relative abundance of the forward and reverse reaction substrates, carnitine and free CoA.

CPT-I and -II are unlikely to be involved because studies of their respective enzyme activities reveal a strong preference for substrates 8 to 16 carbons in length, considerably longer than the C5 chain length of 3MG CoA. On the other hand, CrOT prefers acyl CoA substrates that are 6 to 10 carbons in length, although there are no reports of CrOT accepting terminally carboxylated acyl CoAs. Moreover, despite its apparent acyl CoA substrate compatibility, CrOT is localized to peroxisomes [22,23] and, as such, would appear to be unavailable to catalyze the conversion of mitochondrial 3MG CoA to 3MG carnitine. Thus, the identity of the carnitine acyltransferase enzyme responsible for conversion of 3MG CoA to 3MG carnitine remains unknown at present [24].

5.1. 3MG Carnitine as a Disease Biomarker

Given recent advances in metabolomics and 3MG carnitine detection sensitivity, numerous studies have reported an association between 3MG carnitine and disease [2]. In secondary 3MGC aciduria caused by an IEM in TMEM70, Mackay et al [25] reported the presence of 3MG carnitine along with 3MGC acid and 3MG acid. Moreover, studies have identified 3MG carnitine as a biomarker of several chronic and acute disease pathologies. For example, Kraus et al [26] reported that increased levels of 3MG carnitine in circulation are predictive of cardiovascular disease events. In another study, designed to identify biomarkers of acute coronary syndrome, Wang et al [27] reported elevated levels of 3MG carnitine in patient urine, as compared to healthy controls. Likewise, in a study of subjects with non-ischemic dilated cardiomyopathy, Verdonschot et al [28] reported that 3MG carnitine levels positively correlate with disease severity. In another report, Ruiz et al [29] found that, compared to control subjects, heart failure patients have increased levels of 3MG carnitine. Aside from heart disease, in a study of paraquat-induced kidney toxicity, Wan et al [30] observed that 3MG carnitine levels were 3-fold higher in paraquat-exposed, versus control, subjects. The appearance of 3MG carnitine in other maladies, including traumatic brain injury [31] and following exposure of mice to gamma radiation [32], underscores the potential importance of this metabolite as a biomarker of mitochondrial dysfunction.

Despite reports identifying 3MG carnitine in various metabolomics studies, no consensus has emerged to explain its metabolic origin. As described above, acylcarnitines are formed by carnitine acyltransferase(s) via reversible enzyme reactions between distinct acyl CoA substrates and free carnitine. The metabolic purpose of converting acyl CoAs into their carnitine derivative is twofold: a) to promote the translocation of long-chain fatty acids into mitochondria and b) to export short-chain acyl carnitines out of mitochondria. It may be considered that, when 3MG CoA accumulates in mitochondria, because it is not part of a metabolic pathway, it is converted to 3MG carnitine or 3MG acid. Either of these fates have the dual effect of preserving the mitochondrial pool of free CoA and targeting the metabolically inert 3MG carbon skeleton for export and excretion in urine. It is conceivable that the predominant excretion of 3MG acid in primary and secondary 3MGC acidurias is a result of the relatively large amounts of these acids excreted. This implies that, although formation of the acyl carnitine may be preferred, when carnitine is limiting, more 3MG CoA will be processed to 3MG acid than when carnitine is abundant. Carnitine supplementation studies in subjects with secondary 3MGC aciduria (where 3MG acid is preferably excreted) may provide insight about the apparent discrepancy in 3MG CoA fate in secondary 3MG acidurias versus acute and chronic disease states.

5.2. Can GCDH Mediate Conversion of 3MGC CoA to 3MG CoA?

In considering the metabolic origin of 3MG CoA, as described above, this unique acyl CoA is considered to arise from reduction of the double bond in trans-3MGC CoA. While this is clearly a plausible explanation, the identity of the “reductase” responsible for this reaction has remained a mystery for 40 years. Indeed, given the preponderance of oxidative metabolism in mitochondria, enzyme-mediated reduction of unsaturated acyl CoA substrates is largely unknown. Because 3MG CoA is not an intermediate in any established metabolic pathway, one must consider the possibility that it is formed in a “side reaction”, via a known enzyme that, under specific physiological conditions, accepts trans-3MGC CoA as an alternate substrate.

Based on the strong association between 3MG acid / 3MG carnitine and numerous disease states, deciphering the nature of the enzyme responsible for this reductase activity is of interest. One possible route is a “reverse dehydration” catalyzed by a member of the acyl CoA dehydrogenase (ACAD) enzyme family [33]. Whereas Faull et al [4] proposed that isovaleryl CoA dehydrogenase may be responsible, this suggestion has been challenged [3]. Among the nine members of the acyl CoA dehydrogenase enzyme family, only glutaryl CoA dehydrogenase (GCDH; EC 1.3.99.7) recognizes a terminally carboxylated acyl CoA substrate [34, 35]. In the lysine metabolism pathway, this enzyme catalyzes the flavin-dependent oxidative decarboxylation of glutaryl CoA to crotonyl CoA and CO2. Mutations / deficiency in GCDH are causative for type 1 glutaric aciduria [36, 37]. Herein, it is proposed that, in discrete disease conditions, reduced GCDH (i.e., GCDH-FADH2) accepts trans-3MGC CoA as an alternate substrate and reduces it to 3MG CoA, concomitant with oxidation of its tightly bound flavin prosthetic group. It is further predicted that, in disorders associated with compromised mitochondrial energy metabolism, factors that influence the amount of 3MG CoA generated include the half-life of reduced GCDH and the relative concentrations of glutaryl CoA and trans-3MGC CoA.

Kinetic studies have revealed that GCDH initially catalyzes the oxidation of glutaryl CoA to the unsaturated reaction intermediate, glutaconyl CoA, which remains enzyme bound (Figure 5). In a second step, glutaconyl CoA is decarboxylated to crotonyl CoA and CO2, which are released as products [38]. In the first step, oxidation of glutaryl CoA to glutaconyl CoA coincides with reduction of the FAD prosthetic group. Thus, GCDH contains two distinct catalytic activities; 1) oxidation of the saturated acyl CoA, glutaryl CoA and 2) decarboxylation of the unsaturated, enzyme-bound intermediate, glutaconyl CoA. Compared to other members of the ACAD family, GCDH is unique in that product release and FADH2 oxidation are not coupled [39]. Indeed, studies have revealed that release of crotonyl CoA occurs independent of flavin oxidation state [40]. As with other members of the ACAD family, FADH2 bound to GCDH must be oxidized to FAD before its natural substrate, glutaryl CoA, can bind [33]. Under normal physiological conditions, flavin oxidation is achieved through the action of electron transferring flavoprotein (ETF), which transfers an electron pair from FADH2 to the iron sulfur protein, ETF:ubiquinone oxidoreductase (EC 1.5.5.1). In turn, this oxidoreductase transfers an electron pair to the ETC by reducing coenzyme Q. Thus, ETF plays a crucial role in GCDH activity in vivo because, until it accepts electrons from the enzyme’s reduced flavin, the reaction cycle is not complete. When IEMs, such as those that manifest secondary 3MGC aciduria, adversely affect ETC activity, ETF-dependent oxidation of GCDH’s flavin prosthetic group slows. Under these conditions, it is hypothesized that trans-3MGC CoA serves as substrate for a side reaction (Figure 6), wherein it binds to GCDH-FADH2 at the site formerly occupied by the structurally related molecule, glutaconyl CoA. When this occurs, GCDH-FADH2 reduces the double bond in trans-3MGC CoA, giving rise to 3MG CoA and GCDH-FAD. Because other members of the ACAD family release reaction products only after FADH2 has been oxidized back to FAD by ETF, analogous side reactions cannot occur.

Figure 5. GCDH reaction mechanism under normal physiological conditions.

Figure 5.

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GCDH converts glutaryl CoA to glutaconyl CoA in an oxidation reaction. Glutaconyl CoA is an enzyme-bound intermediate which does not leave the enzyme but, instead, is decarboxylated to form crotonyl CoA and CO2 which dissociate from GCDH as products. During the initial phase of the reaction, GCDH’s flavin prosthetic group is reduced (GCDH-FADH2), and subsequently, donates electrons to the electron transferring flavoprotein (ETF) to become oxidized (GCDH-FAD). Reduced ETF (ETFRed) then donates electrons to ubiquinone oxidoreductase, re-forming ETFOx. Ubiquinone oxidoreductase donates electrons to Complex II of the electron transport chain.

Figure 6. Proposed GCDH-mediated reduction of trans-3MGC CoA to 3MG CoA.

Figure 6.

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As GCDH converts glutaryl CoA into crotonyl CoA and CO2, the enzyme’s flavin prosthetic group is reduced. This FADH2 must be re-oxidized before another glutaryl CoA substrate can bind. Under conditions of detective electron transport chain function, however, this reaction stalls and GCDH-FADH2 accumulates. When this occurs in the presence of trans-3MGC CoA, this metabolite binds to the enzyme at the site previously occupied by glutaconyl CoA and is reduced to 3MG CoA, concomitant with FADH2 oxidation. This reaction cycle can continue as long as glutaryl CoA and trans-3MGC CoA are available.

6. Conclusions

Studies of 3MG acid and 3MG carnitine have revealed that interesting and novel chemical and/or biochemical reactions are responsible for their formation. Both of these molecules originate from 3MG CoA, which is derived from trans-3MGC CoA, a leucine catabolism pathway intermediate. trans-3MGC CoA is a transient intermediate that, normally, is efficiently hydrated by the reversible enzyme, AUH, forming HMG CoA. However, when IEMs block leucine metabolism (primary 3MGC aciduria) or ETC function (secondary 3MGC aciduria), trans-3MGC CoA levels rise. Because it is a labile compound [41], trans-3MGC CoA readily isomerizes to cis-3MGC CoA which spontaneously undergoes intramolecular cyclization to form cis-3MGC anhydride. The anhydride is also reactive and either undergoes hydrolysis to 3MGC acid or it reacts with protein lysine side chain amino groups to form a covalent acyl adduct. Aside from these two fates, the double bond in trans-3MGC CoA can be reduced, forming 3MG CoA. In this review, it is proposed that GCDH is responsible for reduction of trans-3MGC CoA. Once it is formed, 3MG CoA has three potential fates, two non-enzymatic and one enzymatic. The non-enzymatic path is analogous to that of trans-3MGC CoA and involves intramolecular cyclization to form 3MG anhydride followed by either hydrolysis to 3MG acid or covalent acylation of protein lysine side chains. The alternative path is conversion of 3MG CoA to 3MG carnitine. This latter route has taken on additional importance because a growing number of metabolomics studies have shown that various chronic and acute disease process manifest elevated levels of 3MG carnitine [2]. In these disorders, it appears that compromised mitochondrial energy metabolism is a contributing factor. When a given disease manifests mitochondrial dysfunction, the acetyl CoA diversion pathway is activated and trans-3MGC CoA is produced from acetyl CoA. At the same time, as ETC activity slows, the ability to oxidize reduced cofactors, such as FADH2 and NADH, is impeded. One such enzyme is GCDH, which contains a tightly bound FAD prosthetic group. When glutaryl CoA is metabolized via GCDH, GCDH’s flavin group is reduced but, when disease / IEM-related ETC dysfunction exists, the ETF is unable to efficiently oxidize the reduced flavin back to FAD. When this coincides with increased levels of trans-3MGC CoA, conditions are set to generate 3MG CoA. By understanding the molecular basis underlying 3MG carnitine formation, combined with the ability to detect it in biological fluids, this unique short chain dicarboxy acylcarnitine has the potential to serve as a versatile disease prognosticator.

Highlights.

  • 3-methylglutaryl (3MG) CoA is the precursor of 3MG acid and 3MG carnitine

  • The precursor of these waste products is trans-3-methylglutaconyl (3MGC) CoA

  • Reduction of trans-3MGC CoA may involve glutaryl CoA dehydrogenase (GCDH)

  • Mitochondrial dysfunction could cause GCDH to convert trans-3MGC CoA to 3MG CoA

  • Elevated 3MG acid or 3MG carnitine levels represent disease biomarkers

Acknowledgements

This work was supported by a grant from the National Institutes of Health (R37 HL64159) and the Alice and Fred Ottoboni Endowment. The authors thank Irina Romenskaia, Sharon Young, Dr. Kathy Schegg, and Dr. Dylan Jones for contributing to the development of this project.

Abbreviations:

HMG

3-Hydroxy-3-methylglutaryl

3MGC

3-Methylglutaconyl

3MG

3-Methylglutaryl

IEM

Inborn error of metabolism

CAT

Carnitine acyltransferase

AUH

3MGC CoA hydratase

3HIV

3-Hydroxyisovaleric

GCDH

Glutaryl CoA dehydrogenase

ACAD

Acyl CoA dehydrogenase

ETC

Electron transport chain

ETF

Electron transferring flavoprotein

Footnotes

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Conflict of Interest Declaration

The authors have no competing interests to declare.

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