The long chain fatty acyl-CoA ligase (or synthetase) is an

enzyme An enzyme () is a protein that acts as a biological catalyst by accelerating chemical reactions. The molecules upon which enzymes may act are called substrate (chemistry), substrates, and the enzyme converts the substrates into different mol ...

() of the

ligase In biochemistry, a ligase is an enzyme that can catalyze the joining ( ligation) of two molecules by forming a new chemical bond. This is typically via hydrolysis of a small pendant chemical group on one of the molecules, typically resulting i ...

family that activates the oxidation of complex

fatty acids In chemistry, in particular in biochemistry, a fatty acid is a carboxylic acid with an aliphatic chain, which is either saturated or unsaturated. Most naturally occurring fatty acids have an unbranched chain of an even number of carbon atoms, ...

. Long chain fatty acyl-CoA synthetase catalyzes the formation of fatty

acyl-CoA Acyl-CoA is a group of coenzyme A, CoA-based coenzymes that metabolize carboxylic acids. Fatty acyl-CoA's are susceptible to beta oxidation, forming, ultimately, acetyl-CoA. The acetyl-CoA enters the citric acid cycle, eventually forming several e ...

by a two-step process proceeding through an adenylated intermediate. The enzyme catalyzes the following reaction, :

Fatty acid In chemistry, in particular in biochemistry, a fatty acid is a carboxylic acid with an aliphatic chain, which is either saturated and unsaturated compounds#Organic chemistry, saturated or unsaturated. Most naturally occurring fatty acids have an ...

+ CoA + ATP ⇌

Acyl-CoA Acyl-CoA is a group of coenzyme A, CoA-based coenzymes that metabolize carboxylic acids. Fatty acyl-CoA's are susceptible to beta oxidation, forming, ultimately, acetyl-CoA. The acetyl-CoA enters the citric acid cycle, eventually forming several e ...

+ AMP + PP_i It is present in all organisms from bacteria to humans. It catalyzes the pre-step reaction for

β-oxidation In biochemistry and metabolism, beta oxidation (also β-oxidation) is the catabolic process by which fatty acid molecules are broken down in the cytosol in prokaryotes and in the mitochondria in eukaryotes to generate acetyl-CoA. Acetyl-CoA enters ...

of fatty acids or can be incorporated in phospholipids.

Function

Long chain fatty acyl-CoA synthetase, LC-FACS, plays a role in the physiological regulation of various cellular functions via the production of long chain fatty acyl-CoA

esters In chemistry, an ester is a chemical compound, compound derived from an acid (either organic or inorganic) in which the hydrogen atom (H) of at least one acidic hydroxyl group () of that acid is replaced by an organyl group (R). These compounds c ...

, which reportedly have affected

protein Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residue (biochemistry), residues. Proteins perform a vast array of functions within organisms, including Enzyme catalysis, catalysing metab ...

transport, enzyme activation, protein acylation,

cell signaling In biology, cell signaling (cell signalling in British English) is the Biological process, process by which a Cell (biology), cell interacts with itself, other cells, and the environment. Cell signaling is a fundamental property of all Cell (biol ...

, and transcriptional regulation. The formation of fatty acyl-CoA is catalyzed in two steps: a stable intermediate of fatty acyl-AMP molecule and then the product is formed—fatty acid acyl-CoA molecule. Fatty acyl CoA synthetase catalyzes the activation of a long fatty acid chain to a fatty acyl CoA, requiring the energy of 1 ATP to AMP and

pyrophosphate In chemistry, pyrophosphates are phosphorus oxyanions that contain two phosphorus atoms in a linkage. A number of pyrophosphate salts exist, such as disodium pyrophosphate () and tetrasodium pyrophosphate (), among others. Often pyrophosphates a ...

. This step uses 2 "ATP equivalents" because pyrophosphate is cleaved into 2 molecules of inorganic phosphate, breaking a high-energy phosphate bond.

Mechanism and active site

The mechanism for Long Chain Fatty Acyl-CoA Synthetase is a “bi uni uni bi ping-pong” mechanism. The uni and bi prefixes refer to the number of substrates that enter the enzyme and the number of products that leave the enzyme; bi describes a situation where two substrates enter the enzyme at the same time. Ping-pong signifies that a product is released before another substrate can bind to the enzyme. In step one, ATP and a long chain

fatty acid In chemistry, in particular in biochemistry, a fatty acid is a carboxylic acid with an aliphatic chain, which is either saturated and unsaturated compounds#Organic chemistry, saturated or unsaturated. Most naturally occurring fatty acids have an ...

enter the enzyme's

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, the ''binding s ...

. Within the active site the negatively charged oxygen on the fatty acid attacks the alpha phosphate on ATP, forming an ATP-long chain fatty acid intermediate. (Step 1, Figure 2) In the second step,

Pyrophosphate In chemistry, pyrophosphates are phosphorus oxyanions that contain two phosphorus atoms in a linkage. A number of pyrophosphate salts exist, such as disodium pyrophosphate () and tetrasodium pyrophosphate (), among others. Often pyrophosphates a ...

(PPi) leaves, resulting in an AMP-long chain fatty acid molecule within the enzyme's

. (Step 2, Figure 2)

Coenzyme A Coenzyme A (CoA, SHCoA, CoASH) is a coenzyme, notable for its role in the Fatty acid metabolism#Synthesis, synthesis and Fatty acid metabolism#.CE.B2-Oxidation, oxidation of fatty acids, and the oxidation of pyruvic acid, pyruvate in the citric ac ...

now enters the enzyme and another intermediate is formed which consists of AMP-long chain fatty acid-Coenzyme A. (Step 3, Figure 2) At the end of this mechanism two products are released, AMP and acyl coa product. (Step 4, Figure 2) Acyl CoA is formed from long chain fatty acids through an acyl substitution. In an ATP dependent reaction, the fatty acid carboxylate is converted to a

thioester In organic chemistry, thioesters are organosulfur compounds with the molecular structure . They are analogous to carboxylate esters () with the sulfur in the thioester replacing oxygen in the carboxylate ester, as implied by the thio- prefix ...

. The final products of this reaction are

(PPi) and AMP.

Structure

There are several highly conserved areas and a 20-30%

amino acid Amino acids are organic compounds that contain both amino and carboxylic acid functional groups. Although over 500 amino acids exist in nature, by far the most important are the 22 α-amino acids incorporated into proteins. Only these 22 a ...

sequence similarity between the members of this superfamily. The enzymes in the family consist of a large N-terminal and a small C-terminal domain, with the catalytic site positioned between the two domains. Substrate binding may affect the relative positions of the C- and N-terminal domains. The C-terminal domain of LC-FACS is assumed to be in an open conformation when a substrate is absent and in a closed conformation when a

substrate Substrate may refer to: Physical layers *Substrate (biology), the natural environment in which an organism lives, or the surface or medium on which an organism grows or is attached ** Substrate (aquatic environment), the earthy material that exi ...

is bound. The accessibility of the active site to solvent is reduced when the C- and N-terminal domains approach one another. The structure-function relationship between LC-FACS and the formation and processing of the acyl-AMP intermediate was still unclear. A domain swapped dimer is formed by LC-FACS, with

monomer A monomer ( ; ''mono-'', "one" + '' -mer'', "part") is a molecule that can react together with other monomer molecules to form a larger polymer chain or two- or three-dimensional network in a process called polymerization. Classification Chemis ...

interacting at the N-terminal domains. A large electrostatically positive concave is located at the back of the structure in the central valley of the homodimer. Asp15 forms an intermolecular salt bridge with Arg176 in the dimer interactions. An intermolecular hydrogen bond is formed between the main chain

carbonyl In organic chemistry, a carbonyl group is a functional group with the formula , composed of a carbon atom double bond, double-bonded to an oxygen atom, and it is divalent at the C atom. It is common to several classes of organic compounds (such a ...

group of Glu16and the side chain of Arg199. At the interface, Glu175 forms an intermolecular salt bridge with Arg199. The L motif, a six-amino acid peptide linker, connects the large N-terminal domain and a small C-terminal domain of each LC-FACS monomer. The N-terminal domain is composed of two subdomains: a distorted antiparallel

β-barrel In protein structures, a beta barrel (β barrel) is a beta sheet (β sheet) composed of Protein tandem repeats, tandem repeats that twists and coils to form a closed toroidal structure in which the first strand is bonded to the last strand (hydrog ...

and two β-sheets surrounded by α-helices forming an αβαβα sandwich. The small C-terminal globular domain consists of two-stranded β-sheet and a three-stranded antiparallel β-sheet flanked by three α-helices.

Dimer interaction

The dimerization of LC-FACS is stabilized through a

salt bridge In electrochemistry, a salt bridge or ion bridge is an essential laboratory device discovered over 100 years ago. It contains an electrolyte solution, typically an inert solution, used to connect the Redox, oxidation and reduction Half cell, ...

between Asp15 of sequence A and Arg176 of sequence B. Figure 3 shows this salt bridge between these two amino acids. The yellow line between Asp15 and Arg176 shows the salt bridge present.

ATP binding to the C-terminal domain

The conformations of the C-terminal domain of the LC-FACS structures are dependent on the presence of a

ligand In coordination chemistry, a ligand is an ion or molecule with a functional group that binds to a central metal atom to form a coordination complex. The bonding with the metal generally involves formal donation of one or more of the ligand's el ...

. AMP-PNP, a nonhydrolyzable ATP analogue, bound to LC-FACS results in the closed conformation with the C- and N-terminal domains directly interacting. In crystal structures, AMP-PNP is bound in a crevasse of each monomer at the interface between the N- and C-terminal domains. The closed conformation of the C-terminal domain is retained with myristroyl-AMP. Three residues in the C-terminal domain, Glu443, Glu475, and Lys527, interact noncovalently with L motif residues and the N-terminal domain to stabilize the closed conformation. There are two types of open conformations in the C-terminal domains of the uncomplexed structure. The C- and N-terminal domains do not interact directly for both monomers of the dimer. An extensive hydrogen bond network is used by the AMP moiety of the bound ATP molecule to hold the C- and N-terminal domains together.

Fatty acid-binding tunnel

Bulkier long chain fatty acids are bound by a fatty acid-binding tunnel that is located in the N-terminal domain of each

. A large

β-sheet The beta sheet (β-sheet, also β-pleated sheet) is a common structural motif, motif of the regular protein secondary structure. Beta sheets consist of beta strands (β-strands) connected laterally by at least two or three backbone chain, backbon ...

and an

α-helix An alpha helix (or α-helix) is a sequence of amino acids in a protein that are twisted into a coil (a helix). The alpha helix is the most common structural arrangement in the Protein secondary structure, secondary structure of proteins. It is al ...

cluster surround the tunnel which extends from the concave cavity in the central valley to the site of ATP-binding. There are two distinct paths in the large central pathway of the tunnel in the complex structure, which includes the “ATP path” and the “center path,” separated by the indole ring of Trp234 in the G motif. There is also another branch of the central pathway known as the “dead and branch.” The indole ring of Trp234 closes the fatty acid-binding tunnel in the uncomplexed structure. It opens up once AMP-PNP binds through hydrogen bond formation between β-phosphate and the nitrogen on the ring of Trp234. During this time, the closed conformation is adopted by the mobile C-terminal domain. There is a shift in the flexible loop of the G motif in the closed structures of LC-FACS, resulting in a wider dead end branch compared to the uncomplexed forms. The ATP binding site is connected to an ATP path that is a

hydrophobic In chemistry, hydrophobicity is the chemical property of a molecule (called a hydrophobe) that is seemingly repelled from a mass of water. In contrast, hydrophiles are attracted to water. Hydrophobic molecules tend to be nonpolar and, thu ...

channel in the fatty acid-binding tunnel. The fatty acid enters through the center path extending from the interface of the dimer along β-strand 13 to the ATP path. The connection between the two paths is blocked by the indole ring of Trp234 in the absence of ATP. Water molecules fill the center path in the AMP-PNP and myristoyl-AMP complex structures and through the entrance of the center path, they connect to the bulk solvent regions. The basic residues from each monomer, Lys219, Arg296, Arg297, Arg321, Lys350, and Lys 354, cause the entrance of the center path to generate a positive electrostatic potential. The dead end branch contains residues 235-243 and extends from the fatty acid-binding tunnel to α-helix h. The bottom of the dead end branch consists of a hydrophilic environment from the water molecules and polar side chains.

Domains

The domains founds in Long chain fatty acyl CoA synthetase are shown both in the enzyme view (figure 5) and sequence view (figure 6). LC-FACS has five domains. After searching 1v26 in Entrez, the location of the 5 domains was shown and was used to create figure 5 and 6. The ribbons colors in figure 5 correspond to the colors of the figure 6.

Inhibition by long chain fatty acyl-CoAs

A long term and short term regulation controls fatty acid synthesis. Long term fatty acid synthesis regulation is dependent on the rate of acetyl-CoA carboxylase (ACC) synthesis, the rate-limiting enzyme and first enzyme of the fatty acid synthesis, and fatty acid synthase (FAS), the second and major enzyme of the fatty acid synthesis. Cellular fatty acyl-CoA is involved in the short term regulation, but there is not a full understanding of the mechanisms. Free fatty acids inhibits the de novo fatty acid synthesis and appears to be dependent on the formation of long chain fatty acyl-CoAs. Studies have shown that long chain fatty acyl-CoAs inhibit ACC and FAS via feedback inhibition. Long chain fatty acyl-CoA's inhibitory effect on the fatty acid synthesis may be a result of its regulation of lipogenic enzymes in a feedback manner through gene transcription suppression. Long-chain fatty-acid-CoA ligase in cells catalytically synthesizes long chain fatty acyl-CoAs. Long-chain fatty-acid-CoA ligase may be involved in an important role in the suppression of fatty acid synthesis and it has been reported that it played a part in fatty acid synthesis inhibition. It was recently found that vitamin D₃ upregulates FACL3, which forms long-chain fatty acid synthesis through the use of

myristic acid Myristic acid (IUPAC name: tetradecanoic acid) is a common saturated fatty acid with the molecular formula . Its salts and esters are commonly referred to as myristates or tetradecanoates. The name of the acyl group derived from myristic acid is m ...

eicosapentaenoic acid Eicosapentaenoic acid (EPA; also icosapentaenoic acid) is an omega−3 fatty acid. In physiological literature, it is given the name 20:5(''n''−3). It also has the trivial name timnodonic acid. In chemical structure, EPA is a carboxylic acid wi ...

(EPA), and

arachidonic acid Arachidonic acid (AA, sometimes ARA) is a polyunsaturated omega−6 fatty acid 20:4(ω−6), or 20:4(5,8,11,14). It is a precursor in the formation of leukotrienes, prostaglandins, and thromboxanes. Together with omega−3 fatty acids an ...

as substrates, in expression and activity levels. FACL3 contributes to vitamin D₃ growth inhibitory effect in human prostate cancer LNCaP cells. A current study reports that the feedback inhibition of FAS expression by long chain fatty acyl-CoAs causes the downregulation of FAS mRNA by vitamin D₃.

Clinical significance

Adrenoleukodystrophy (ALD) is the build up of long chain fatty acids in the brain and adrenal cortex, because of the decreased activity of long chain fatty acyl coa synthetase. The

oxidation Redox ( , , reduction–oxidation or oxidation–reduction) is a type of chemical reaction in which the oxidation states of the reactants change. Oxidation is the loss of electrons or an increase in the oxidation state, while reduction is ...

of the long chain fatty acids normally occurs in the peroxisome where the long chain fatty acyl coa synthetase is found. Long chain fatty acids enter the peroxisome via a transporter protein, ALDP, which creates a gate in the membrane of the

peroxisome A peroxisome () is a membrane-bound organelle, a type of microbody, found in the cytoplasm of virtually all eukaryotic cells. Peroxisomes are oxidative organelles. Frequently, molecular oxygen serves as a co-substrate, from which hydrogen perox ...

. In ALD the gene for this peroximal membrane transporter, ALDP, is defective, preventing long chain fatty acids from entering the peroxisome.

Examples

Human genes encoding long-chain-fatty-acid—CoA ligase enzymes (also known as acyl-CoA synthetase long-chain, or ACSL) include: * ACSL1 * ACSL3 * ACSL4 * ACSL5 * ACSL6 * SLC27A2

References

External links

* {{DEFAULTSORT:Long-chain-fatty-acid-CoA ligase EC 6.2.1 Beta oxidation Oncogenes