Direct Evidence for the Size and Conformational Variability of the Pyruvate Dehydrogenase Complex Revealed by Three-dimensional Electron Microscopy

Zhou, Zheng; Liao, Wangcai; Cheng, R. Holland; Lawson, Janet E.; McCarthy, Diane; Reed, Lester J.; Stoops, James K

doi:10.1074/jbc.m101765200

Cited by 54 publications

(58 citation statements)

References 46 publications

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“…The finding of a premature stop codon in the a13 allele strongly suggests that the mutant phenotype is caused by defective Dlat. In addition, the Leu-Pro substitution at position 644 in the m631 allele is likely to disrupt core formation and PDH activity, because the C terminus of Dlat is intimately involved in forming intertrimer contacts (30).…”

Section: Resultsmentioning

confidence: 99%

A zebrafish model for pyruvate dehydrogenase deficiency: Rescue of neurological dysfunction and embryonic lethality using a ketogenic diet

Taylor

Hurley

Epps

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

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Defects in the pyruvate dehydrogenase (PDH) complex result in severe neurological dysfunction, congenital lactic acidosis, growth retardation, and early death. Current treatments for PDH deficiency are administered postnatally and are generally unsuccessful. Because many patients with this disease are born with irreversible defects, a model system for the development of effective pre-and postnatal therapies would be of great value. In a behavioral genetic screen aimed to identify zebrafish with visual function defects, we previously isolated two alleles of the recessive lethal mutant no optokinetic response a (noa). Here we report that noa is deficient for dihydrolipoamide S-acetyltransferase (Dlat), the PDH E2 subunit, and exhibits phenotypes similar to human patients with PDH deficiency. To rescue the deficiency, we added ketogenic substrates to the water in which the embryos develop. This treatment successfully restored vision, promoted feeding behavior, reduced lactic acidosis, and increased survival. Our study demonstrates an approach for establishing effective therapies for PDH deficiency and other congenital diseases that affect early embryonic development.H uman diseases that affect tissues with high-energy requirements are often caused by defects in mitochondrial function (1). Pyruvate dehydrogenase (PDH) deficiency is a rare metabolic disorder that severely affects the central nervous system due to the high-energy demand of neurons and often results in developmental delay, feeding difficulties, lethargy, ataxia, blindness, and early death (2-4). PDH is a nuclear-encoded mitochondrial matrix multienzyme complex that provides the primary link between glycolysis and the tricarboxylic acid (TCA) cycle by catalyzing the irreversible conversion of pyruvate into acetyl-CoA (5). Defects in PDH result in decreased acetyl-CoA synthesis and the accumulation of pyruvate and lactate, causing reduced energy production and metabolic acidosis, respectively.Current treatments for PDH deficiency, which include the activation of residual PDH with dichloroacetate (6), the administration of cofactors (lipoic acid and thiamine) (7,8), and the provision of ketogenic diets (9, 10), have yielded only limited or variable success (see Fig. 1). The technical challenges of working with small numbers of human patients has made it difficult to include all of the necessary controls and to systematically evaluate therapeutic agents and diets with varying proportions of calories from fat, carbohydrate, and protein (9, 11). Furthermore, preexisting neurological disorders compound the difficulty of evaluating treatments. A therapy that bypasses the entire complex would potentially be of greatest benefit, because it could be used to treat patients with defects in any of the PDH subunits and varying degrees of deficiency.An animal model for PDH deficiency would be of great value to improve current treatments and to allow the development of novel therapies. This model should exhibit phenotypic characteristics similar to those seen in human ...

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Section: Resultsmentioning

confidence: 99%

A zebrafish model for pyruvate dehydrogenase deficiency: Rescue of neurological dysfunction and embryonic lethality using a ketogenic diet

Taylor

Hurley

Epps

et al. 2004

Proc. Natl. Acad. Sci. U.S.A.

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“…Moreover, the 250-Å-diameter dodecahedron has a very unusual feature: the tightly bound trimers at each of its 20 vertices seem to be interconnected by 30 flexible bridges enabling the core to ''breathe,'' as evidenced by an extraordinary size variability of 40 Å (17%) at room temperature. The breathing core apparently is a common feature in the phylogeny of the PDCs, suggesting that protein dynamics is an integral component of the function of these multienzyme complexes (5). Moreover, dodecahedral morphology of the core favors a synchronous or harmonious change in the length of the bridges that is related to the size variation and the function of the complex (5).…”

mentioning

confidence: 99%

“…The breathing core apparently is a common feature in the phylogeny of the PDCs, suggesting that protein dynamics is an integral component of the function of these multienzyme complexes (5). Moreover, dodecahedral morphology of the core favors a synchronous or harmonious change in the length of the bridges that is related to the size variation and the function of the complex (5).…”

mentioning

confidence: 99%

The remarkable structural and functional organization of the eukaryotic pyruvate dehydrogenase complexes

Zhou¹,

McCarthy²,

O’Connor³

et al. 2001

Proc. Natl. Acad. Sci. U.S.A.

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The three-dimensional reconstruction of the bovine kidney pyruvate dehydrogenase complex (M r Ϸ 7.8 ؋ 10 6 ) comprising about 22 molecules of pyruvate dehydrogenase (E 1) and about 6 molecules of dihydrolipoamide dehydrogenase (E 3) with its binding protein associated with the 60-subunit dihydrolipoamide acetyltransferase (E2) core provides considerable insight into the structural and functional organization of the largest multienzyme complex known. The structure shows that potentially 60 centers for acetylCoA synthesis are organized in sets of three at each of the 20 vertices of the pentagonal dodecahedral core. These centers consist of three E 1 molecules bound to one E2 trimer adjacent to an E3 molecule in each of 12 pentagonal openings. The E1 components are anchored to the E 1-binding domain of the E2 subunits through an Ϸ50-Å-long linker. Three of these linkers emanate from the outside edges of the triangular base of the E 2 trimer and form a cage around its base that may shelter the lipoyl domains and the E 1 and E2 active sites. The docking of the atomic structures of E1 and the E 1 binding and lipoyl domains of E2 in the electron microscopy map gives a good fit and indicates that the E 1 active site is Ϸ95 Å above the base of the trimer. We propose that the lipoyl domains and its tether (swinging arm) rotate about the E 1-binding domain of E 2, which is centrally located 45-50 Å from the E1, E2, and E3 active sites, and that the highly flexible breathing core augments the transfer of intermediates between active sites. T he pyruvate dehydrogenase complex (PDC) serves as the link between glycolysis and the tricarboxylic acid cycle and generally has prominence in the description of these metabolic pathways because they serve as a major source of cellular energy. A central feature of PDCs is a 24-mer (Escherichia coli) or 60-mer (eukaryotes and some Gram-positive bacteria) core with the morphologies of a cube or pentagonal dodecahedron, respectively (1-4). The structures with the latter morphology comprise the largest (M r Ϸ 10 7 ) multienzyme complexes known. Even more remarkable than their exceptional size and morphology, these complexes encompass some of the most unusual features found in structural biology as described below.The E 2 core comprises the dihydrolipoamide acetyltransferase activity and is the only oligomeric enzyme complex known to be organized with the shape of a pentagonal dodecahedron. Moreover, the 250-Å-diameter dodecahedron has a very unusual feature: the tightly bound trimers at each of its 20 vertices seem to be interconnected by 30 flexible bridges enabling the core to ''breathe,'' as evidenced by an extraordinary size variability of 40 Å (17%) at room temperature. The breathing core apparently is a common feature in the phylogeny of the PDCs, suggesting that protein dynamics is an integral component of the function of these multienzyme complexes (5). Moreover, dodecahedral morphology of the core favors a synchronous or harmonious change in the length of the bridges that is relate...

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“…However, the structures of the E2 core and entire PDC from yeast and mammals were studied by cryoelectron microscopy (Fig. 4) [17,18]. Both electron microscopy and crystal structures of the E2 core revealed the trimer building blocks in the cubic (8 trimers) or dodecahedral (20 trimers) structures.…”

Section: Structure and Organization Of Pdcmentioning

confidence: 99%

“…The contraction and expansion of the E2 core was proposed to reflect the conformational mobility of PDC during Three serine residues of E1 phosphorylation sites are shown as S1, S2, and S3 in dephosphorylated condition and as P1, P2, and P3 when phosphorylated. catalysis [17]. Recently the structure of the whole PDC from bovine kidney was constructed based on cryoelectron microscopy ( Fig.…”

Section: Structure and Organization Of Pdcmentioning

confidence: 99%

The biochemistry of the pyruvate dehydrogenase complex*

Patel

Korotchkina

2003

Biochem Molecular Bio Educ

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In the Westernized world the daily dietary caloric requirements are roughly provided as follows: 40% from carbohydrates, 40% from fats, and 20% from proteins. In some populations in developing countries the daily caloric contribution from carbohydrate is even higher due to its readily available sources and relatively low cost. Glucose, the principal product of carbohydrate digestion, passes through a series of enzymatic steps first in the non-oxidative glycolytic pathway (from glucose to pyruvate) followed by efficient oxidative metabolism via the tricarboxylic acid cycle (from acetyl-CoA to CO 2 and H 2 O) to harness a portion of its potential energy as ATP. Interestingly, these two major pathways are directly linked by the pyruvate dehydrogenase complex (PDC) 1 localized in the mitochondrial matrix (Fig. 1). In the fed state acetyl-CoA generated from pyruvate (derived mostly from glucose and some dietary amino acids) is also utilized for the biosynthesis of lipids such as long chain fatty acids and cholesterol by lipogenic tissues (such as liver and adipose tissues and under special conditions in mammary glands during lactation and in the brain during the prenatal and early postnatal development). Additionally, amino acids from excess dietary protein are metabolized by several specialized reactions or pathways generating intermediates that have to be ultimately converted to pyruvate first and then to acetylCoA via PDC either for complete oxidation to CO 2 and H 2 O or for lipogenesis. PDC is the only known reaction in most eukaryotes to generate acetyl-CoA (two-carbon compound) from pyruvate (three-carbon compound). Since this is a physiologically irreversible reaction and since there is no other known reaction or pathway to convert the two-carbon compound to pyruvate (or its equivalent) for the synthesis of glucose in animals, the flux through PDC is tightly regulated to meet the specific metabolic and energetic needs of different tissues during the fed and fasting (starvation) states. This is accomplished by covalent modification of the ratelimiting component of the complex involving sophisticated interplay among the components of the complex and allosteric modulations by acetyl-CoA and NADH, the products of the reaction (and also of fatty acid oxidation). It is evident from these simple considerations that PDC plays a key role as a gatekeeper of both caloric and glucose homeostasis in mammals. In this review, we will discuss recent developments concerning the structure-function relationship of this multienzyme complex from various organisms with emphasis on regulatory aspects of the mammalian complex. Detailed accounts of various aspects of this complex can be found in several excellent reviews [1][2][3][4][5][6][7][8][9][10][11]. STRUCTURE AND ORGANIZATION OF PDCPDC is present in most prokaryotic and eukaryotic organisms. It catalyzes several sequential reactions of oxidative decarboxylation of pyruvic acid by the action of its three catalytic components: (i) pyruvate dehydrogenase (E1) catalyzing the de...

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Direct Evidence for the Size and Conformational Variability of the Pyruvate Dehydrogenase Complex Revealed by Three-dimensional Electron Microscopy

Cited by 54 publications

References 46 publications

A zebrafish model for pyruvate dehydrogenase deficiency: Rescue of neurological dysfunction and embryonic lethality using a ketogenic diet

A zebrafish model for pyruvate dehydrogenase deficiency: Rescue of neurological dysfunction and embryonic lethality using a ketogenic diet

The remarkable structural and functional organization of the eukaryotic pyruvate dehydrogenase complexes

The biochemistry of the pyruvate dehydrogenase complex*

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