How phosphorylation influences E1 subunit pyruvate dehydrogenase: A computational study

Sgrignani, Jacopo; Chen, Jing Jing; Alimonti, Andrea; Cavalli, Andrea

doi:10.1038/s41598-018-33048-z

Cited by 19 publications

(25 citation statements)

References 70 publications

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“…PDH is one of the three enzymatic subunits (E1, E2, and E3) forming the pyruvate dehydrogenase complex (PDC) that catalyzes the conversion of pyruvate in Acetyl-CoA. Particularly, PDH (corresponding to E1 subunit) catalyzed pyruvate decarboxylation which is the first step in the conversion of pyruvate in Acetyl-CoA [ 22 ]. It has been reported that PDH activity is greatly decreased in cachectic conditions [ 23 ].…”

Section: Resultsmentioning

confidence: 99%

A Metabolic Change towards Fermentation Drives Cancer Cachexia in Myotubes

et al. 2021

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Cachexia is a disorder associated with several pathologies, including cancer. In this paper, we describe how cachexia is induced in myotubes by a metabolic shift towards fermentation, and the block of this metabolic modification prevents the onset of the cachectic phenotype. Cachectic myotubes, obtained by the treatment with conditioned medium from murine colon carcinoma cells CT26, show increased glucose uptake, decreased oxygen consumption, altered mitochondria, and increased lactate production. Interestingly, the block of glycolysis by 2-deoxy-glucose or lactate dehydrogenase inhibition by oxamate prevents the induction of cachexia, thus suggesting that this metabolic change is greatly involved in cachexia activation. The treatment with 2-deoxy-glucose or oxamate induces positive effects also in mitochondria, where mitochondrial membrane potential and pyruvate dehydrogenase activity became similar to control myotubes. Moreover, in myotubes treated with interleukin-6, cachectic phenotype is associated with a fermentative metabolism, and the inhibition of lactate dehydrogenase by oxamate prevents cachectic features. The same results have been achieved by treating myotubes with conditioned media from human colon HCT116 and human pancreatic MIAPaCa-2 cancer cell lines, thus showing that what has been observed with murine-conditioned media is a wide phenomenon. These findings demonstrate that cachexia induction in myotubes is linked with a metabolic shift towards fermentation, and inhibition of lactate formation impedes cachexia and highlights lactate dehydrogenase as a possible new tool for counteracting the onset of this pathology.

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Section: Resultsmentioning

confidence: 99%

A Metabolic Change towards Fermentation Drives Cancer Cachexia in Myotubes

et al. 2021

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“…Phosphorylation of Ser232 is seen to behave very differently compared to Ser293 and Ser300, with the phosphorylation level increasing even during lactate consumption. [36] used a molecular dynamics method and could not identify inhibitory effects due to pSer232 in the cases where pSer293 and pSer300 inhibited enzyme activity [36]. Although in vitro studies have shown that the phosphorylation of any of the three Ser residues on the E1

α

subunit of PDC leads to the deactivation of the enzyme complex [35], pSer293 was found to have the most potent inhibitory effect [58, 59].…”

Section: Resultsmentioning

confidence: 99%

“…Further insights into the highly dynamic nature of PDC as a multienzyme complex could be achieved using mathematical modeling approaches. [36] used molecular dynamic simulations to test the structural consequences of the phosphorylation on

PDC E 1 α

. They found that phosphorylations of Ser293 and Ser300 change the

PDC E 1 α

catalytic site [36].…”

Section: Introductionmentioning

confidence: 99%

“…[36] used molecular dynamic simulations to test the structural consequences of the phosphorylation on

PDC E 1 α

. They found that phosphorylations of Ser293 and Ser300 change the

PDC E 1 α

catalytic site [36]. As a different approach, [22] developed an in vitro and in vivo kinetic model of the PDC reaction network and [23] used the same model to study the influence of pyruvate concentration and suggested that the PDC reaction rate is a non‐linear function of its substrate.…”

Section: Introductionmentioning

confidence: 99%

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Regulation of pyruvate dehydrogenase complex related to lactate switch in CHO cells

Möller

Bhat

Guhl

et al. 2020

Engineering in Life Sciences

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The metabolism of Chinese hamster ovary (CHO) cell lines is typically characterized by high rates of aerobic glycolysis with increased lactate formation, known as the ”Warburg” effect. Although this metabolic state can switch to lactate consumption, the involved regulations of the central metabolism have only been partially studied so far. An important reaction transferring the lactate precursor, pyruvate, into the tricarboxylic acid cycle is the decarboxylation reaction catalyzed by the pyruvate dehydrogenase enzyme complex (PDC). Among other mechanisms, PDC is mainly regulated by phosphorylation–dephosphorylation at the three sites Ser232, Ser293, and Ser300. In this work, the PDC phosphorylation in antibody‐producing CHO DP‐12 cell culture is investigated during the lactate switch. Batch cultivations were carried out with frequent sampling (every 6 h) during the transition from lactate formation to lactate uptake, and the PDC phosphorylation levels were quantified using a novel indirect flow cytometry protocol. Contrary to the expected activation of PDC (i.e., reduced PDC phosphorylation) during lactate consumption, Ser293 and Ser300 phosphorylation levels were 33% higher compared to the phase of glucose excess. At the same time, the relative phosphorylation level of Ser232 increased steadily throughout the cultivation (66% increase overall). The intracellular pyruvate was found to accumulate only during the period of high lactate production, while acetyl‐CoA showed nearly no accumulation. These results indicate a deactivation of PDC and reduced oxidative metabolism during lactate switch even though the cells undergo a metabolic transition to lactate‐based cell growth and metabolism. Overall, this study provides a unique view on the regulation of PDC during the lactate switch, which contributes to an improved understanding of PDC and its interaction with the bioprocess.

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“…PDC activity is mainly regulated via the phosphorylation of the E1 component with regulation independent of phosphorylation also reported. 11,31–34 Inactivation of the E1 component of human PDC is achieved via phosphorylation of E1α S203, S264, and S271 located on loops formed by E1α amino acid stretches R259-R282 (Loop A, highly conserved) and N195-E205 (Loop B, highly conserved) (Figure 1). 11,31,32,35,36 E1α S264 is the most rapidly phosphorylated site.…”

Section: Introductionmentioning

confidence: 99%

Simulations of Pathogenic E1α Variants: Allostery and Impact on Pyruvate Dehydrogenase Complex-E1 Structure and Function

Gökcan

Bedoyan

Isayev

2022

Preprint

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Pyruvate dehydrogenase complex (PDC) deficiency is a major cause of primary lactic acidemia resulting in high morbidity and mortality, with limited therapeutic options. The E1 component of the mitochondrial multienzyme PDC (PDC-E1) is a symmetric dimer of heterodimers (αβ/α’β’) encoded by the PDHA1 and PDHB genes, with two symmetric active sites each consisting of highly conserved phosphorylation loops A and B. PDHA1 mutations are responsible for 82-88% of cases. Greater than 85% of E1α residues with disease-causing missense mutations (DMMs) are solvent inaccessible, with ~30% among those involved in subunit-subunit interface contact (SSIC). We performed molecular dynamics simulations of wild-type (WT) PDC-E1 and E1 variants with E1α DMMs at R349 and W185 (residues involved in SSIC), to investigate their impact on human PDC-E1 structure. We evaluated the change in E1 structure and dynamics and examined their implications on E1 function with the specific DMMs. We found that the dynamics of phosphorylation Loop A which is crucial for E1 biological activity, changes with DMMs that are at least about 15 Å away. Because communication is essential for PDC-E1 activity (with alternating active sites), we also investigated the possible communication network within WT PDC-E1 via centrality analysis. We observed that DMMs altered/disrupted the communication network of PDC-E1. Collectively, these results indicate allosteric effect in PDC-E1, with implications for development of novel small molecule therapeutics for specific recurrent E1α DMMs such as replacements of R349 responsible for ~10% of PDC deficiency due to E1α DMMs.

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How phosphorylation influences E1 subunit pyruvate dehydrogenase: A computational study

Cited by 19 publications

References 70 publications

A Metabolic Change towards Fermentation Drives Cancer Cachexia in Myotubes

A Metabolic Change towards Fermentation Drives Cancer Cachexia in Myotubes

Regulation of pyruvate dehydrogenase complex related to lactate switch in CHO cells

Simulations of Pathogenic E1α Variants: Allostery and Impact on Pyruvate Dehydrogenase Complex-E1 Structure and Function

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