Boyer — Academic Press, Wellen, K. ATP-citrate lyase links cellular metabolism to histone acetylation. Science , — Sun, T. Identification of the citrate-binding site of human ATP-citrate lyase using X-ray crystallography. Fan, F. On the catalytic mechanism of human ATP citrate lyase.
Biochemistry 51 , — Wei, X. Aoshima, M. A novel enzyme, citryl-CoA lyase, catalysing the second step of the citrate cleavage reaction in Hydrogenobacter thermophilus TK Inoue, H. Studies on ATP citrate lyase of rat liver. The role of CoA. Biochem 65 , — Watson, J. Tricarballylate and hydroxycitrate: substrate and inhibitor of ATP: citrate oxaloacetate lyase. Rokita, S. Biochemistry 21 , — Download references. This project was supported by a grant from Research Foundation Flanders to K.
G0G and a Ghent University grant to S. In the 's-early 's, four German-born biochemists, Fritz Lipmann, Hans Krebs, Feodor Lynen and Konrad Bloch, were investigating the mechanism by which glucose foods are metabolised in the body and turned into either fats for storage, or energy for immediate use.
They worked independently, since three of them had fled from the Nazis to seek refuge in countries like Switzerland, the UK or US. Fritz Lipmann "for his discovery of co-enzyme A and its importance for intermediary metabolism". Hans Krebs "for his discovery of the citric acid cycle". Feodor Lynen Konrad Bloch "for their discoveries concerning the mechanism and regulation of the cholesterol and fatty acid metabolism". Between them, they found that one of the central molecules involved in this process is a coenzyme a molecule that helps an enzyme , which was named coenzyme A or CoA for short.
The A stood for acetyl, since one of CoA's main jobs is to transfer two-carbon units in the form of acetyl between various biological molecules. It can be thought of as the body's 'delivery truck', since it transports its cargo of C 2 along the roadways of blood vessels to the retail stores muscles where it's unloaded.
CoA is composed of two main parts, a long protein-like chain shown in black in the figure , joined to adenosine diphosphate, ADP, shown in blue which is one of the molecules along with ATP used for energy storage.
The important part of the molecule is at the end of the protein chain, which terminates in a sulph-hydryl -SH group red. This group is highly reactive, and links to carboxylic acid molecules via a thioester bond. The most important acid is acetic acid, and when it is joined to CoA, the resulting compound is known as acetyl-CoA. However, all of these two-carbon acetyl units are in the wrong place. Before they can be used in fatty acid synthesis, they have to be moved into the cytoplasm of the cell, where the fatty acids will be made.
Acetyl-CoA is moved through the mitochondrial membrane, and enters the cytoplasm of the cell, as the molecule citrate. In the cytoplasm, these citrate molecules are once again converted back to acetyl-CoA.
This reaction requires that the cell use up some energy by breaking down an ATP molecule. Fatty acids are made by repeatedly joining together the two-carbon fragments found in acetyl-CoA and then reducing the -CO- part of the molecule to -CH 2 -. In this way, the hydrocarbon chain, which will become the hydrophobic, energy storing part of the fatty acid, grows two-carbons at a time as the cycle of joining reactions is repeated over and over again.
Most of these reactions take place in and on the membranes of the endoplasmic reticulum known as microsomal membranes and takes place in several stages. In step three, an acetyl-group is attached to the cysteine-SH carrier site on the FAS complex and a malonyl-group is attached to the pantethine-SH carrier site.
Fatty acids are a significant source of this mitochondrial acetyl-CoA pool [ 13 ]. As a result, acetyl-CoA is generated in the mitochondria for oxidation or other possible fates. Under such conditions, lower nucleocytosolic acetyl-CoA will also limit fatty acid synthesis, histone acetylation, and other growth-related processes. ATP citrate lyase is inhibited under these situations at both the transcriptional and post-translational levels [ 17 , 18 ].
Depletion of nucleocytosolic acetyl-CoA also represents a cue to induce autophagy [ 19 , 20 ]. More generally, many other genes with functions involved in stress and survival are induced concomitantly with core autophagy genes [ 21 , 23 , 24 ]. These genes tend to be less dependent on histone acetylation for their activation [ 11 , 25 , 26 ], perhaps due to reduced availability of acetyl-CoA.
In mammalian cells, autophagy regulation by acetyl-CoA occurs in a manner dependent on the p acetyltransferase [ 20 ]. Thus, the regulation of autophagy by acetyl-CoA may occur primarily at the level of transcriptional control of core autophagy genes.
Taken together, under fasted or carbon-poor states, nucleocytosolic amounts of acetyl-CoA decrease in cells, while mechanisms to channel acetyl-CoA into the mitochondria are engaged. These considerations support a model in which the subcellular compartmentalization of acetyl-CoA units undergoes a major shift during starvation, and the utilization of these acetyl units is re-purposed to support survival strategies Fig.
How might cells actually sense the abundance of acetyl-CoA? It is perhaps no coincidence that acetyl-CoA doubles as the acetyl donor for protein acetylation modifications including histone acetylation Fig.
Studies performed under carbon-rich conditions where acetyl-CoA synthesis is not limiting may mask the contributions of this metabolite in cellular regulation. However, most organisms, as well as particular tissue microenvironments in vivo experience challenges in the nutrient environment that might limit acetyl-CoA biosynthesis or availability e.
Recent studies have begun to provide compelling evidence that many protein acetylation modifications are indeed modulated by acetyl-CoA availability [ 27 , 28 ]. A The acetylation of proteins may be catalyzed by acetyltransferase enzymes or can occur spontaneously through reaction with acetyl-CoA directly.
Deacetylase enzymes catalyze the removal of acetylation modifications. Liberated acetate can be converted back to acetyl-CoA. The removal of aberrant acetylation or acylation modifications may restore protein function.
Besides histones, the acetyl-CoA synthetase family of enzymes was also identified to be regulated by reversible acetylation [ 29 — 31 ]. The acetylation of an active site lysine residue was observed to inhibit the activity of acetyl-CoA synthetase as a mechanism of feedback inhibition in response to high acetyl-CoA [ 32 — 34 ].
The deacetylation of these enzymes, catalyzed by sirtuins, restores their activity [ 32 — 34 ]. Subsequent mass spectrometry surveys have now revealed that thousands of other proteins, including many other metabolic enzymes, can be acetylated [ 35 — 38 ].
In some cases, every enzyme in a particular biochemical pathway was found to be acetylated [ 39 ]. Although the majority of these modifications were found to be inhibitory, several were reported to be activating [ 40 ].
In some instances, the acetylation of particular metabolic enzymes was responsive to glucose levels in the media, suggesting that they could be linked to intracellular acetyl-CoA abundance. Whether specific acetyltransferase enzymes catalyze the majority of these acetylation modifications present on metabolic enzymes is not yet clear.
The yeast metabolic cycle YMC offers a system to investigate whether particular acetylation modifications might be coupled to acetyl-CoA itself. Studies of yeast cells undergoing the YMC during continuous, glucose-limited growth in a chemostat have revealed periodic changes in intracellular acetyl-CoA amounts as yeast cells alternate between growth and quiescent-like phases [ 22 ]. Several proteins are dynamically acetylated precisely in phase with the observed acetyl-CoA oscillations [ 11 ].
Interestingly, the dynamic acetylation of all of these proteins is dependent on the acetyltransferase Gcn5p, suggesting this enzyme has the capability of acetylating its substrates in tune with acetyl-CoA fluctuations in vivo. Moreover, acetylation of SAGA subunits appears to aid its recruitment to growth genes [ 11 ]. A brief survey of other acetylated proteins that are not known to be Gcn5p substrates showed they are not dynamically acetylated across the YMC [ 11 ].
An analysis of the genomic regions bound by these acetylated histones revealed that several marks, in particular H3K9Ac, were present predominantly at growth genes, specifically during the growth phase of the YMC when acetyl-CoA levels rise [ 11 , 25 ].
These considerations suggest that the acetylation of these nuclear-localized proteins collectively functions to promote the activation of growth genes in response to a burst of nucleocytosolic acetyl-CoA. Given the thousands of newly identified acetylated proteins, a pertinent question is what proportion of each protein is acetylated? Recent studies aiming to determine the stoichiometry of acetylated sites estimate that for many proteins, only a small fraction of the peptides are actually acetylated [ 42 , 43 ].
However, nuclear proteins, including histones and transcription factors, were estimated to be acetylated at much higher stoichiometry [ 43 ]. Conventional shotgun detection of peptides by mass spectrometry is biased towards abundant proteins, so perhaps it is unsurprising that a small fraction of a very abundant protein that is acetylated could be scored as a positive.
Moreover, lysine residues on proteins can react spontaneously with thioesters such as acetyl-CoA or other acyl-CoA metabolites, resulting in non-enzymatic acetylation or acylation [ 44 — 48 ]. Non-enzymatic acetylation or acylation may be especially prominent within the mitochondria [ 43 , 46 , 48 , 49 ], which is thought to have higher acetyl-CoA concentrations and higher pH, thereby increasing the nucleophilicity of lysyl side chains.
Thus, while some non-enzymatic acetylation or acylation events could have evolved to be regulatory, the possibility also exists that many of these modifications could be spurious. These considerations must be taken into account when determining the physiological significance of any detected acetylation site. Moreover, there are limitations to mutation of a lysine residue to either arginine or glutamine. These mutations are not always accurate acetylated or deacetylated lysine mimics, and could perturb protein function independent of site-specific acetylation.
As such, it can be challenging to demonstrate whether a particular acetylation modification is functionally important in vivo. To help address these issues, methods for site-specific incorporation of acetyllysine [ 50 ], as well as better acetylated or deacetylated lysine mimics, have been developed [ 51 , 52 ]. The use of these and other methods will help clarify the extent through which protein acetylation modifications are responsive to acetyl-CoA fluctuations in a regulatory manner, either enzymatically or non-enzymatically.
The accumulation of acetyl-CoA in subcellular compartments may also necessitate the activity of deacetylase enzymes to remove non-enzymatic acetylation modifications that could intentionally or unintentionally compromise protein function [ 28 , 53 , 54 ].
Consistent with this idea, hyperacetylation of mitochondrial enzymes occurs in the absence of mitochondrial SIRT3 [ 55 — 57 ], and deacetylation of these enzymes typically increases their activity [ 53 ]. Moreover, the expression of SIRT3 is increased specifically under fasting states, in response to high-fat diets, or during exercise - conditions that all promote increased mitochondrial acetyl-CoA [ 53 ].
Likewise, the potential of proteins to be modified by other acyl-CoA metabolites besides acetyl-CoA is supported by the discovery of a wide variety of acylation modifications present on proteins, along with associated sirtuins that preferentially catalyze their removal [ 58 — 61 ].
Evidence that sirtuins evolved specifically to remove non-enzymatic protein acylation as a form of protein quality control has been summarized in a recent review [ 54 ]. In this model, failure of sirtuins to remove aberrant acylation modifications would hinder the function of effected proteins and consequently lead to dysfunctions in metabolism and susceptibility to disease [ 47 , 55 , 57 ].
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