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must be bound by the enzyme to cause a conformational change in the
Acetyl-CoA synthetase (ACS) or Acetate—CoA ligase is an
+ enzyme
Enzymes () are proteins that act as biological catalysts by accelerating chemical reactions. The molecules upon which enzymes may act are called substrates, and the enzyme converts the substrates into different molecules known as products. A ...
() involved in metabolism of acetate. It is in the ligase
In biochemistry, a ligase is an enzyme that can catalyze the joining (ligation) of two large molecules by forming a new chemical bond. This is typically via hydrolysis of a small pendant chemical group on one of the larger molecules or the enzym ...
class of enzymes, meaning that it catalyzes the formation of a new chemical bond between two large molecules.
Reaction
The two molecules joined together that make up Acetyl CoA areacetate
An acetate is a salt (chemistry), salt formed by the combination of acetic acid with a base (e.g. Alkali metal, alkaline, Alkaline earth metal, earthy, Transition metal, metallic, nonmetallic or radical Radical (chemistry), base). "Acetate" als ...
and coenzyme A
Coenzyme A (CoA, SHCoA, CoASH) is a coenzyme, notable for its role in the synthesis and oxidation of fatty acids, and the oxidation of pyruvate in the citric acid cycle. All genomes sequenced to date encode enzymes that use coenzyme A as a subs ...
(CoA). The complete reaction with all the substrates and products included is:
: ATP + Acetate + CoA <=> AMP #REDIRECT Amp #REDIRECT Amp
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{{Redirect category shell, {{R from other capitalisation{{R from ambiguous page ...Pyrophosphate
In chemistry, pyrophosphates are phosphorus oxyanions that contain two phosphorus atoms in a P–O–P linkage. A number of pyrophosphate salts exist, such as disodium pyrophosphate (Na2H2P2O7) and tetrasodium pyrophosphate (Na4P2O7), among other ...
+ Acetyl-CoA
Acetyl-CoA (acetyl coenzyme A) is a molecule that participates in many biochemical reactions in protein, carbohydrate and lipid metabolism. Its main function is to deliver the acetyl group to the citric acid cycle (Krebs cycle) to be oxidized for ...
Once acetyl-CoA is formed it can be used in the TCA cycle
The citric acid cycle (CAC)—also known as the Krebs cycle or the TCA cycle (tricarboxylic acid cycle)—is a series of chemical reactions to release stored energy through the oxidation of acetyl-CoA derived from carbohydrates, fats, and proteins ...
in aerobic respiration to produce energy and electron carriers. This is an alternate method to starting the cycle, as the more common way is producing acetyl-CoA from pyruvate
Pyruvic acid (CH3COCOOH) is the simplest of the alpha-keto acids, with a carboxylic acid and a ketone functional group. Pyruvate, the conjugate base, CH3COCOO−, is an intermediate in several metabolic pathways throughout the cell.
Pyruvic aci ...
through the pyruvate dehydrogenase complex
Pyruvate dehydrogenase complex (PDC) is a complex of three enzymes that converts pyruvate into acetyl-CoA by a process called pyruvate decarboxylation. Acetyl-CoA may then be used in the citric acid cycle to carry out cellular respiration, and t ...
. The enzyme's activity takes place in the mitochondrial matrix
In the mitochondrion, the matrix is the space within the inner membrane. The word "matrix" stems from the fact that this space is viscous, compared to the relatively aqueous cytoplasm. The mitochondrial matrix contains the mitochondrial DNA, ribo ...
so that the products are in the proper place to be used in the following metabolic steps. Acetyl Co-A can also be used in fatty acid synthesis
In biochemistry, fatty acid synthesis is the creation of fatty acids from acetyl-CoA and NADPH through the action of enzymes called fatty acid synthases. This process takes place in the cytoplasm of the cell. Most of the acetyl-CoA which is conve ...
, and a common function of the synthetase is to produce acetyl Co-A for this purpose.
The reaction catalyzed by acetyl-CoA synthetase takes place in two steps. First, AMP #REDIRECT Amp #REDIRECT Amp
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{{Redirect category shell, {{R from other capitalisation{{R from ambiguous page ...active site
In biology and biochemistry, the active site is the region of an enzyme where substrate molecules bind and undergo a chemical reaction. The active site consists of amino acid residues that form temporary bonds with the substrate (binding site) a ...
, which allows the reaction to take place. The active site is referred to as the A-cluster. A crucial lysine
Lysine (symbol Lys or K) is an α-amino acid that is a precursor to many proteins. It contains an α-amino group (which is in the protonated form under biological conditions), an α-carboxylic acid group (which is in the deprotonated −C ...
residue must be present in the active site to catalyze the first reaction where Co-A is bound. Co-A then rotates in the active site into the position where acetate can covalently bind to CoA. The covalent bond is formed between the sulfur atom in Co-A and the central carbon atom of acetate.
The ACS1 form of acetyl-CoA synthetase is encoded by the gene facA, which is activated by acetate and deactivated by glucose.
Structure
The three dimensional structure of the asymmetric ACS (RCSB PDB ID number: 1PG3) reveals that it is composed of two subunits. Each subunit is then composed primarily of two domains. The larger N-terminal domain is composed of 517 residues, while the smaller C-terminal domain is composed of 130 residues.; Each subunit has an active site
In biology and biochemistry, the active site is the region of an enzyme where substrate molecules bind and undergo a chemical reaction. The active site consists of amino acid residues that form temporary bonds with the substrate (binding site) a ...
where the ligands are held. The crystallized
Crystallization is the process by which solid forms, where the atoms or molecules are highly organized into a structure known as a crystal. Some ways by which crystals form are precipitating from a solution, freezing, or more rarely depos ...
structure of ACS was determined with CoA and Adenosine- 5′-propylphosphate bound to the enzyme. The reason for using Adenosine- 5′-propylphosphate is that it is an ATP competitive inhibitor
Competitive inhibition is interruption of a chemical pathway owing to one chemical substance inhibiting the effect of another by competing with it for binding or bonding. Any metabolic or chemical messenger system can potentially be affected b ...
which prevents any conformational changes to the enzyme. The adenine ring of AMP/ATP is held in a hydrophobic pocket create by residues Ile (512) and Trp (413).
The source for the crystallized structure is the organism Salmonella typhimurium (strain LT2 / SGSC1412 / ATCC 700720). The gene for ACS was then transfected into Escherichia coli BL21(DE3) for expression. During chromatography in the process to isolate the enzyme, the subunits came out individually and the total structure was determined separately. The method used to determine the structure was X-ray diffraction
X-ray crystallography is the experimental science determining the atomic and molecular structure of a crystal, in which the crystalline structure causes a beam of incident X-rays to diffract into many specific directions. By measuring the angles ...
with a resolution of 2.3 angstroms. The unit cell values and angles are provided in the following table:
Function
The role of the ACS enzyme is to combine acetate and CoA to form acetyl CoA, however its significance is much larger. The most well known function of the product from this enzymatic reaction is the use ofAcetyl-CoA
Acetyl-CoA (acetyl coenzyme A) is a molecule that participates in many biochemical reactions in protein, carbohydrate and lipid metabolism. Its main function is to deliver the acetyl group to the citric acid cycle (Krebs cycle) to be oxidized for ...
in the role of the TCA cycle
The citric acid cycle (CAC)—also known as the Krebs cycle or the TCA cycle (tricarboxylic acid cycle)—is a series of chemical reactions to release stored energy through the oxidation of acetyl-CoA derived from carbohydrates, fats, and proteins ...
as well as in the production of fatty acid. This enzyme is vital to the action of histone acetylation
Histone acetyltransferases (HATs) are enzymes that acetylate conserved lysine amino acids on histone proteins by transferring an acetyl group from acetyl-CoA to form ε-''N''-acetyllysine. DNA is wrapped around histones, and, by transferring an ...
as well as gene regulation. The effect this acetylation has is far reaching in mammals. It has been shown that downregulation of the acs gene in the hippocampal
The hippocampus (via Latin from Greek , 'seahorse') is a major component of the brain of humans and other vertebrates. Humans and other mammals have two hippocampi, one in each side of the brain. The hippocampus is part of the limbic system, an ...
region of mice results in lower levels of histone acetylation, but also impairs the long-term spatial memory of the animal. This result points to a link between cellular metabolism, gene regulation and cognitive function. This enzyme has shown to be an interesting biomarker for the presence of tumors in colorectal carcinomas. When the gene is present, the cells are able to take in acetate as a food source to convert it to Acetyl-CoA during stressed conditions. In the cases of advanced carcinoma tumors, the genes for this enzyme were down regulated and indicated a poor 5-year survival rate. Expression of the enzyme has also been linked to the development of metastatic tumor nodes, leading to a poor survival rate in patients with renal cell carcinomas.
Regulation
The activity of the enzyme is controlled in several ways. The essentiallysine
Lysine (symbol Lys or K) is an α-amino acid that is a precursor to many proteins. It contains an α-amino group (which is in the protonated form under biological conditions), an α-carboxylic acid group (which is in the deprotonated −C ...
residue in the active site plays an important role in regulation of activity. The lysine molecule can be deacetylated by another class of enzyme called sirtuins
Sirtuins are a family of signaling proteins involved in metabolic regulation. They are ancient in animal evolution and appear to possess a highly conserved structure throughout all kingdoms of life. Chemically, sirtuins are a class of proteins ...
. In mammals, the cytoplasmic-nuclear synthetase (AceCS1) is activated by SIRT1
Sirtuin 1, also known as NAD-dependent deacetylase sirtuin-1, is a protein that in humans is encoded by the SIRT1 gene.
SIRT1 stands for sirtuin (silent mating type information regulation 2 homolog) 1 (''S. cerevisiae''), referring to the fact t ...
while the mitochondrial synthetase (AceCS2) is activated by SIRT3
NAD-dependent deacetylase sirtuin-3, mitochondrial also known as SIRT3 is a protein that in humans is encoded by the ''SIRT3'' gene irtuin (silent mating type information regulation 2 homolog) 3 (S. cerevisiae) SIRT3 is member of the mammalian sir ...
. This action increases activity of this enzyme. The exact location of the lysine residue varies between species, occurring at Lys-642 in humans, but is always present in the active site of the enzyme.
Since there is an essential allosteric
In biochemistry, allosteric regulation (or allosteric control) is the regulation of an enzyme by binding an effector molecule at a site other than the enzyme's active site.
The site to which the effector binds is termed the ''allosteric site ...
change that occurs with the binding of an AMP molecule, the presence of AMP can contribute to regulation of the enzyme. Concentration of AMP must be high enough so that it can bind in the allosteric binding site and allow the other substrates to enter the active site. Also, copper ions deactivate acetyl Co-A synthetase by occupying the proximal site of the A-cluster active site, which prevents the enzyme from accepting a methyl group to participate in the Wood-Ljungdahl Pathway.
The presence of all the reactants in the proper concentration is also needed for proper functioning as in all enzymes.
Acetyl-CoA synthetase is also produced when it is needed for fatty acid synthesis
In biochemistry, fatty acid synthesis is the creation of fatty acids from acetyl-CoA and NADPH through the action of enzymes called fatty acid synthases. This process takes place in the cytoplasm of the cell. Most of the acetyl-CoA which is conve ...
, but, under normal conditions, the gene is inactive and has certain transcriptional factors that activate transcription when necessary.
In addition to sirtuins, protein deacetylase (AcuC) also can modify acetyl-CoA synthetase at a lysine residue. However, unlike sirtuins, AcuC does not require NAD+ as a cosubstrate.
Role in gene expression
While acetyl-CoA synthetase's activity is usually associated with metabolic pathways, the enzyme also participates in gene expression. In yeast, acetyl-CoA synthetase delivers acetyl-CoA to histone acetyltransferases for histone acetylation. Without correct acetylation, DNA cannot condense intochromatin
Chromatin is a complex of DNA and protein found in eukaryotic cells. The primary function is to package long DNA molecules into more compact, denser structures. This prevents the strands from becoming tangled and also plays important roles in r ...
properly, which inevitably results in transcriptional errors.
Industrial application
By taking advantage of the pathways which use acetyl-CoA as a substrate, engineered products can be obtained which have potential to be consumer products. By overexpressing the acs gene, and using acetate as a feedstock, the production of fatty acids (FAs) may be increased. The use of acetate as a feed stock is uncommon, as acetate is a normal waste product of ''E. coli'' metabolism and is toxic at high levels to the organism. By adapting the E. coli to use acetate as a feedstock, these microbes were able to survive and produce their engineered products. These fatty acids could then be used as a biofuel after being separated from the media, requiring further processing (transesterification
In organic chemistry, transesterification is the process of exchanging the organic group R″ of an ester with the organic group R' of an alcohol. These reactions are often catalyzed by the addition of an acid or base catalyst. The reaction can ...
) to yield usable biodiesel fuel. Original adaptation protocol for inducing high levels of acetate uptake was innovated in 1959 as a means to induce starvation mechanisms in ''E. coli''.
Intracellular
Acetyl-CoA from the breakdown of sugars in glycolysis have been used to build fatty acids. However the difference comes in the fact that the Keasling strain is able to synthesize its own ethanol, and process ( by transesterification) the fatty acid further to create stable fatty acid ethyl esters (FAEEs). Removing the need for further processing prior to obtaining a usable fuel product in Diesel engines.
''Acetyl CoA used in the production of both ethanol and fatty acids''
Trans-esterification
Preliminary studies have been conducted where the combination of these two methods have resulted in the production of FAEEs, using acetate as the only carbon source using a combination of the methods described above. The levels of production of all methods mentioned are not up to levels required for large scale applications (yet).References
{{DEFAULTSORT:Acetyl-CoA synthetase EC 6.2.1