The alterations in myocardial energy substrate metabolism that occur in heart failure, and the causes and consequences of these abnormalities, are poorly understood. There is evidence to suggest th...

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Myocardial Substrate Metabolism in the Normal and Failing Heart

First conceptualized as a mechanism for the mitochondrial transport of long-chain fatty acids in the early 1960s, the carnitine palmitoyltransferase (CPT) system has since come to be recognized as a pivotal component of fuel homeostasis. This is by virtue of the unique sensitivity of the outer membrane CPT I to the simple molecule, malonyl-CoA. In addition, both CPT I and the inner membrane enzyme, CPT II, have proved to be loci of inherited defects, some with disastrous consequences. Early efforts using classical approaches to characterize the CPT proteins in terms of structure/function/regulatory relationships gave rise to confusion and protracted debate. By contrast, recent application of molecular biological tools has brought major enlightenment at an exponential pace. Here we review some key developments of the last 20 years that have led to our current understanding of the physiology of the CPT system, the structure of the CPT isoforms, the chromosomal localization of their respective genes, and the identification of mutations in the human population.

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1997.00001.x

The mitochondrial carnitine palmitoyltransferase system. From concept to molecular analysis.

Readers of this journal do not need to be reminded of the explosive increase in obesity/type 2 diabetes syndromes and their attendant staggering public health costs that are currently afflicting developed countries. Because of this alarming development and what it portends for the future if left unchecked, the American Diabetes Association, together with its counterparts around the world, as well as the World Health Organization, are faced with the daunting challenge of finding answers to two huge questions. First, what is the pathophysiological basis for these disorders? Second, why is their rate of appearance accelerating so rapidly at this particular juncture of human history, and what, if anything, can we do to intervene? The primary focus of this lecture will be on the first of these issues. I want to emphasize at the outset that I do not plan to dwell exclusively on our own studies in this area. Instead, I shall try to incorporate recent developments from a variety of other laboratories working in the field. Although these will of necessity have to be highly selective, I hope that collectively they will illustrate how we have come to a relatively new and exciting way of thinking about the etiology of type 2 diabetes.

It has been said in literary circles that James Joyce’s Ulysses is probably the most frequently opened and least read book that has ever been published. In a similar vein, I sometimes think that the question of what causes type 2 diabetes might be one of the most frequently asked and least satisfactorily answered in the history of diabetes research. This is not meant to be an arrogant statement, nor indeed does it imply that I have a magical solution to the question at hand. Rather, it simply reflects the fact that at the dawn of …

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Banting Lecture 2001: Dysregulation of Fatty Acid Metabolism in the Etiology of Type 2 Diabetes

A single entity, the AMP-activated protein kinase (AMPK), phosphorylates and regulates in vivo hydroxymethylglutaryl-CoA reductase and acetyl-CoA carboxylase (key regulatory enzymes of sterol synthesis and fatty acid synthesis, respectively), and probably many additional targets. The kinase is activated by high AMP and low ATP via a complex mechanism, which involves allosteric regulation, promotion of phosphorylation by an upstream protein kinase (AMPK kinase), and inhibition of dephosphorylation. This protein-kinase cascade represents a sensitive system, which is activated by cellular stresses that deplete ATP, and thus acts like a cellular fuel gauge. Our central hypothesis is that, when it detects a 'low-fuel' situation, it protects the cell by switching off ATP-consuming pathways (e.g. fatty acid synthesis and sterol synthesis) and switching on alternative pathways for ATP generation (e.g. fatty acid oxidation). Native AMP-activated protein kinase is a heterotrimer consisting of a catalytic alpha subunit, and beta and gamma subunits, which are also essential for activity. All three subunits have homologues in budding yeast, which are components of the SNF1 protein-kinase complex. SNF1 is activated by glucose starvation (which in yeast leads to ATP depletion) and genetic studies have shown that it is involved in derepression of glucose-repressed genes. This raises the intriguing possibility that AMPK may regulate gene expression in mammals. AMPK/SNF1 homologues are found in higher plants, and this protein-kinase cascade appears to be an ancient system which evolved to protect cells against the effects of nutritional or environmental stress.

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1997.00259.x

The AMP‐Activated Protein Kinase

5-Aminoimidazole-4-carboxamide ribonucleoside (AICAR) has previously been reported to be taken up into cells and phosphorylated to form ZMP, an analog of 5′-AMP This study was designed to determin

AICA riboside increases AMP-activated protein kinase, fatty acid oxidation, and glucose uptake in rat muscle

The requirement for carnitine and the malonyl-CoA sensitivity of carnitine palmitoyl-transferase I (EC 2.3.1.21) were measured in isolated mitochondria from eight tissues of animal or human origin using fixed concentrations of palmitoyl-CoA (50 microM) and albumin (147 microM). The Km for carnitine spanned a 20-fold range, rising from about 35 microM in adult rat and human foetal liver to 700 microM in dog heart. Intermediate values of increasing magnitude were found for rat heart, guinea pig liver and skeletal muscle of rat, dog and man. Conversely, the concentration of malonyl-CoA required for 50% suppression of enzyme activity fell from the region of 2-3 microM in human and rat liver to only 20 nM in tissues displaying the highest Km for carnitine. Thus, the requirement for carnitine and sensitivity to malonyl-CoA appeared to be inversely related. The Km of carnitine palmitoyltransferase I for palmitoyl-CoA was similar in tissues showing large differences in requirement for carnitine. Other experiments established that, in addition to liver, heart and skeletal muscle of fed rats contain significant quantities of malonyl-CoA and that in all three tissues the level falls with starvation. Although its intracellular location in heart and skeletal muscle is not known, the possibility is raised that malonyl-CoA (or a related compound) could, under certain circumstances, interact with carnitine palmitoyltransferase I in non-hepatic tissues and thereby exert control over long chain fatty acid oxidation.

/pdf/observations-on-the-affinity-for-carnitine-and-malonyl-coa-4fthru5fcv.pdf

Observations on the affinity for carnitine, and malonyl-CoA sensitivity, of carnitine palmitoyltransferase I in animal and human tissues. Demonstration of the presence of malonyl-CoA in non-hepatic tissues of the rat.

The sensitivity of carnitine palmitoyltransferase I (CPT I; EC 2.3.1.21) to inhibition by malonyl-CoA and related compounds was examined in isolated mitochondria from liver, heart and skeletal muscle of the rat. In all three tissues the same order of inhibitory potency emerged: malonyl-CoA much greater than succinyl-CoA greater than methylmalonyl-CoA much greater than propionyl-CoA greater than acetyl-CoA. For any given agent, suppression of CPT I activity was much greater in skeletal muscle than in liver, with the heart enzyme having intermediate sensitivity. With skeletal-muscle mitochondria a high-affinity binding site for [14C]malonyl-CoA was readily demonstrable (Kd approx. 25 nM). The ability of other CoA esters to compete with [14C]malonyl-CoA for binding to the membrane paralleled their capacity to inhibit CPT I. Palmitoyl-CoA also competitively inhibited [14C]malonyl-CoA binding, in keeping with its known ability to overcome malonyl-CoA suppression of CPT I. For reasons not yet clear, free CoA displayed anomalous behaviour in that its competition for [14C]malonyl-CoA binding was disproportionately greater than its inhibition of CPT I. Three major conclusions are drawn. First, malonyl-CoA is not the only physiological compound capable of suppressing CPT I, since chemically related compounds, known to exist in cells, also share this property, particularly in tissues where the enzyme shows the greatest sensitivity to malonyl-CoA. Second, malonyl-CoA and its analogues appear to interact with the same site on the mitochondrial membrane, as may palmitoyl-CoA. Third, the degree of site occupancy by inhibitors governs the activity of CPT I.

Interaction of malonyl-CoA and related compounds with mitochondria from different rat tissues. Relationship between ligand binding and inhibition of carnitine palmitoyltransferase I.

The kinetics of carnitine palmitoyltransferase I (CPT I; EC 2.3.1.21) were examined in mitochondria from rat liver, heart and skeletal muscle as a function of pH over the range 6.8-7.6. In all three tissues raising the pH resulted in a fall in the Km for carnitine, no change in the Km for palmitoyl-CoA or Octanoyl-CoA, and a marked decrease in the inhibitory potency of malonyl-CoA. Studies with skeletal-muscle mitochondria established that increasing pH was accompanied by an increase in the Kd of the malonyl-CoA binding site for this ligand, coupled with a decrease in the Kd for fatty acyl-CoA species to compete for malonyl-CoA binding. Three principal conclusions are drawn. (1) The pH-induced shift in malonyl-CoA sensitivity of CPT I is not a phenomenon restricted to liver mitochondria. (2) At any given pH within the range tested, the ability of malonyl-CoA (and closely related compounds) to inhibit enzyme activity is governed by the efficiency of their binding to the malonyl-CoA site. (3) The competitive interaction between fatty acyl-CoA substrates and malonyl-CoA as regards CPT I activity is exerted at the malonyl-CoA binding site. Finally, the possibility is strengthened that the malonyl-CoA binding site is distinct from the active site of CPT I.

S. E. Mills

Papers

Observations on the affinity for carnitine, and malonyl-CoA sensitivity, of carnitine palmitoyltransferase I in animal and human tissues. Demonstration of the presence of malonyl-CoA in non-hepatic tissues of the rat.

Interaction of malonyl-CoA and related compounds with mitochondria from different rat tissues. Relationship between ligand binding and inhibition of carnitine palmitoyltransferase I.

Effects of pH on the interaction of substrates and malonyl-CoA with mitochondrial carnitine palmitoyltransferase I.

Interaction of malonyl-CoA and 2-tetradecylglycidyl-CoA with mitochondrial carnitine palmitoyltransferase I.

Interaction of substrates and malonyl-CoA with mitochondrial carnitine palmitoyltransferase I