Enzymes are biocatalysts formed in cells either as simple or as conjugated proteins with a non-amino acid component.

Coenzymes often function in the transfer of electrons or of functional groups (hydrogen atom, acetyl, methyl, amino groups, etc.). The coenzymes are generally identical with vitamins which, at least in higher organisms, represent essential components of their food that cannot be synthesized in their organs. The enzymes accelerate biological reactions by decreasing the activation energy of a given reaction without altering its equilibrium. The mechanism of their action consists in the formation of a complex between enzyme and substrate (ES) which undergoes the chemical reaction proper whereupon the enzyme-product complex (EP) is split to the original enzyme and the product. The existence of some enzymesubstrate complexes has now been proved both indirectly (spetrophotometry) and directly (chemical isolation). The substrate is bound to the enzyme in the active site or centre which may be visualized as a spatial arrangement of certain amino acids of the protein moiety of the enzyme as well as of the prosthetic group or the coenzyme. Amino acids frequently found to play a role in the active site are serine (through its OH group) and histidine (through the nitrogen of its imidazole). Some enzymes occur in the active form directly upon their synthesis while others are synthesized as inactive proenzymes (pepsinogen, trypsinogen) or exist a part of their life-time in an inactive conformation (allosteric enzymes). They must then be activated by a special process to become functional. Enzyme-catalyzed reactions proceed at different rates, depending on (a) the amount or activity of the enzyme, (b) the concentration of substrate, (c) the pH and composition of the solution, (d) temperature, (e) the presence of activators and inhibitors. a. Since the enzyme concentrations in living cells are difficult to estimate we often speak about their activities. Enzyme activity is measured in international units (U), corresponding to an activity converting 1 (1 substrate per min, or, more recently, in katals (kat), corresponding to aiactivity converting 1 mol substrate per s. Specific activity of an isolated enzyme is expressed in units per mg protein. Specificity of an enzyme defines the range of structure types the enzyme can attack. If the specificity is absolute the enzyme can catalyze the chemical reaction of a single compound, if it is relative, several related compounds can serve as substrate. Stereospecificity indicates that the enzyme accepts only a certain stereo-isomer (L- or D-form). A given substrate can be converted by one of several theoretically possible reactions catalyzed by different enzymes. This is important in some metabolic control processes. b. At low substrate concentrations the enzyme reaction follows first-order kinetics, i.e. the rate is proportional to substrate concentration. At very high concentrations of substrate the reaction is of zero order, i.e. the enzyme is saturated by its substrate. Every enzyme reaction has its characteristic Michaelis constant, , which defines the concentration at which the enzyme reaction proceeds at half its maximum rate. It is not identical with the dissociation constant of the ES complex just as its reciprocal is not identical with the association constant of this complex. Every enzyme-catalyzed reaction has its pH optimum. d. Every enzyme-catalyzed reaction proceeds most rapidly at a certain temperature. e. Activators and inhibitors are compounds that can bear on the rate of the enzyme reaction either positively or negatively. Activators increase the enzyme reaction by aiding the formation of a functional active site and are most often represented by metal ions, such as Mg2+, Zn2+, Mn2+, Co2+. Activation of allosteric enzymes which are composed of subunits is of paramount importance in the control of multi-enzyme systems; this is usually achieved by various organic molecules. Enzyme inhibitors can be of several kinds, the most important being competitive and noncompetitive ones. Competitive inhibitors interact with the active site of the enzyme and resemble the substrate in its structure so that they compete with it for binding. They may be removed by excess substrate. Noncompetitive inhibitors react with another important structure of the enzyme molecule (e.g., a SH group). The inhibition can be either reversible or irreversible in the case that the inhibitor brings about a permanent (covalent) change of an essential functional group of the enzyme. A noncompetitive inhibitor cannot be removed by excess substrate. To distinguish between the various types of inhibition a number of kinetic and graphical techniques can be used (e.g., that of Lineweaver and Burk). Under in vivo conditions (in cells) most enzymes are spatially organized in the so-called multi-enzyme systems. These are either bound to cell structures or are freely dissolved in various cell compartments. Their intracellular concentrations are usually higher than those of the available intermediates (millimolar with most metabolites). Enzymes are named after the reaction they catalyze, by attaching the suffix -ase, and are divided into six main classes. Both systematic and trivial enzyme names are in use.

~ , Turnover number

mol converted substrate =----------------------------------mm enzyme activity catalyzing the _ conversion of 1 jxmol substrate min number of katals = ----------:----------- kg active protein number of katals = ----------.-------------mol enzyme

EXPRESSIONS EN Z Y M O L O G Y

USED IN

International enzyme unit U _ .Specific activity ,, , Molar activity

A new unit of enzyme activity was introduced in 1973 under the name katal (kat), corresponding to the amount of catalyst able to convert 1 mol substrate to its product per second. The international enzyme unit of activity U is related to the katal as follows: 1 kat = 1 mol s"1 = 60 mol min-1 = 60 x 106 Hinol min-1 = 6 x 107U 1 U = 1 ymol min-1 - (1/60) 1 s-1 = (1/60) (Aat = 16.67 nkat

M E C H A N I S M OF E N Z Y M E A C T I O M In enzyme-catalyzed reactions a transient enzymesubstrate complex is formed. The substrate is bound to the enzyme at the active site which is formed by a definite spatial conformation of the peptide chain(s) where the side chains of some amino acids (Ser, His Tyr, and others) are brought close to each other. The substrate is bound to the active site at three points, the complex formed being usually unstable and breaking down to the product and the free enzyme after the chemical reaction proper. The side chains of amino acids contributing to the active site are usually ionizable (acid), nucleophi-lic (electrondonating) or electrophilic (electron-attracting). The nucleophilic groups may be replaced with metal ions which, in some cases, form an integral part of some enzymes (Mn2+, Mg2+, Zn2+). All the groups participate in the binding of the substrate and in its conversion to the product.

;:>;'''

.- \-

'. ' v 7 '. -v

In some enzymes, the active site conformation is partly due to the molecule of a coenzyme, such as shown here for the pyridoxal phosphate bound in the active site of an enzyme. Interaction of the amino group (Glu) with the carbonyl group of pyridoxal phosphate gives rise to a Schiff base (an aldimine bond). The structure of the protein part of the enzyme molecule assists in the stabilitation of the meso-meric structures derived from the Schiff base. Depending on the nature of the protein molecule, the amino acid can be transaminated, decarboxylated or isomerized.

Nature of group transferred Free Bond on coenzyme

Name of coenzyme

Name of enzyme (code number) Hexokinase (2.7.1.1)

Function (examples of coenzyme dependent rections) Phosphate transfer (to and from OR.) Hexoses phosphorylation Pyrophosphate transfer Phosphorylation of D-riboso-5-phosphate to 5-phosphonbo syl-1 -pyrophosphate Adenylate transfer (to and from RCO) ll Amino acids activation

ATP

-"

ATP

Ri bosophosphate pyrophosphate kinase (pyrophospho transferase) (2 7.16) ArmnoacyJ-tRNA , synthetases (Itgases) (6.1.1.4 to 6.1 1.21)

-"-

ATP

-"-

ATP GTP

Methyl adenosyl transferase (2.5.1 6) Phosphoenol pyruvate carboxy kinase (4.1.1.32)

Adenosyl transfer to and from methionine Formation of phosphoenol pyruvate from oxatoacetate (Some phosphate transfer reactions.) Some carbohydrate i ntercon verstons (mannose -+ fucose)

GDP

Mannose 1phosphate guanilyl tranferase (2 7.7.13)

ITP.

Phosphoenol pyruvate carboxy kinase (41 1 32)

See GTP

UDP

Uridyltransferases (2.7.7.9 to 2.7.712)

Carbohydrate mterconversions, oxidations, isomerisations, synthesis

CTP

Cyti dy .transferases Phospholipid (2.781, 2.7.714; 2.7 7 intercon versions 15)

NADH

Dehydrogenases (pyridine linked) (111.1 to 11.1.45)

Reversible transfer of two reducing equivalents from substrate to the coenzyme oxidized form Carrier of energy-rich electrons. Transfer from catabolic reactions to electron requiring anabolic reactions Synthesis of some hydrogen -rich biomolecufes Direct transfer of two hydrogen atoms from substrate to the oxidized form of coenzyme (oxidative deamination of amino acids) See FADH2 Transfer of "active aldehyde" (glycoialdehyde, acetaldehyde) and dihydroxyetny! group

NADPH

Dehydrogenases (pyridine linked) (1.1.1.10; 1.1119; 1 1 1 34; 1.1 1 36; 1.11.49)

/ADH2

Dehydrogenases (flavin linked) (1.3.99.1, 1.3.99.2)

FMNH, TPP (thiamine pyrophosphate)

Dehyd rogenases (fiavine linked) Tranai do lase (2.2.1 2) TransketoUse (2 2.1 1)

Pyridoxal phosphate

Transaminase aminotransferase) (2.6.1.1. to 2.6.1-52) decarboxylase amino acid) [4.1.1.22) acemase

Transformation of ct-L-amino acids to: a-ketoaci ds amines oc-D-amino acids

a) b) )

=0 ,

Biotin (Ncarboxylate)

Carboxylase (6.4.1.1 to 6.4.1.3)

Carbon-dioxld transfer. [Pyruvate carboxylase, acetyl CoA carboxylase, propionyl CoA carboxylase)

N10 formy! FH* 10~formyl-5. 6, 7, 8tetrahydro folic acid

Phosphoribosyi amino-Imidazolcarboxamid formyl transferase (2.1.2.3)

Formyl transfer. Biosynthesis of purine nucleotides

N* formyl FH4 5formyl-5, 6. 7, otetrahydrofoltc acid

For my (transferase (2.1.2.6).

Formyl transfer. Biosynthesis of purine nucleotides

N**10methenyl FH* 5,10-methenyl-5, 6, 7,8-tetrahydrofollc acid

Phosphori bosy I -glyeinamid formyl transferase (2.1.2.2)

Formyl transfer. Biosynthesis of purine nucleotides

N5 formimino FbU 5formimino-5, 6, 7, 8tetrahydrofolic add

Glutamate formimino transferase (2.1.2.5)

Transfer of formimino group from amino acids (Gly, Glu)

N5*10 methylen FH 5,IO-methylen-5. 6. 7,8-tetra-hydrofolic acid

Thy m idyl ate synthetase

Methyfation of dUMP todTMP

N* methyl FH4 5methyl-5, 6, 7, 8-tetrahydrofolie acid

Methyltransferase

Methyfation of homocysteine to methionine

Co bam ide

Methylmalonyl-CoA mutase (5.4.99.2)

Rearrangement and synthesis of methyl group. Isomer is ation of methylmalonyl CoA to succinyl CoA, glutamic acid to ^-methylaspartic aci

CoA-SH Coenzyme A

Acetyltransferase (2.3.1.1 to 2.3.1.12) Acyltransferase (2.3.1.15 . to 2.3.1.20)

Reactions of carboxylic acids (including acetate). Biosynthesis of carboxylic acids

S-aeyl li poate reduced

Pyruvate dehydrogenase (1.2.4.1) Ketogluurate dehydrogenase (1.2.4.2)

Transfer of ! residues generated by a-keto acids decarboxylation

PAPS Phosphoadenosine phosphosulfate

Sulfotransferase (2.8.2.1 to 2.8.2.5)

Transfer of sulfate to phenol, steroids, arylamine, chondroitine

Coenzyme Pyridoxal phosphate

Function Transamination Decarboxylation Raoemization Aerobic decarboxylation Transfer of the aldehyde group Transfer of acyls Aerobic degradation and synthesis of fatty acids Transfer of carbon groups Transfer of C02 Transfer of H+ + eTransfer of H+ + eTransfer of H+ + eTransfer of H+ + e-

Corresponding vitamin Pyridoxine (Be)

COENZYMES AM i N S

A N I? R E L A T Si V ST

Thiamine pyrophosphate

Thiamine (Bi)

Coenzyme A

Pantothenic acid

It is one of the characteristics of coenzymes that higher organisms cannot synthesize them and, therefore, they must be supplied with food. Because their function is solely catalytic the daily requirement is low (several mg per day in man). Compounds known as vitamins in human diet are in most cases identical with or closely related to coenzymes. This fact underlies the importance of vitamins in nutrition physiology. If vitamins are not in ample supply, metabolic disturbances may occur, known as hypovitaminoses and avitaminoses.

Tetrahydrofolic acid Biotin NAD + NADP+ FMN FAD

Folic acid Biotin (H) Nicotinic acid (niacin) () Nicotinic acid (PP) Riboflavin (Ba) Riboflavin ()

E F F E C T OF pH ON E N Z Y M E ACTIVITY

Most enzymes are characterized by their pH dependence, there being a certain pH value of optimum enzyme activity. On both sides of this value the activity is lower. The optimum pH values range widely from highly acidic (pepsin) to highly alkaline (alkaline phosphatase). Therefore, in all enzyme studies, the pH must be maintained by a suitable buffer. The pH dependence of enzyme activity is determined by the pK of ionizable groups of the enzyme molecule, particularly those at or near the active site (possibly playing a role in the binding of the coenzyme), and those that can contribute to-changes of the active site through conformational changes of parts of the protein molecules. The pH can further affect the degree of ionization or the spatial organization of substrate (including proteins). Most pronounced are the differences in the pH optimum in the digestive tract.
E F F E C T OF T E M P E R A T U R E ON ~ N! X Y H ACTIVITY

Temperature always affects the reaction rate. Within the physiological range, the reaction rate will increase with temperature (1) but above a certain point enzyme-catalyzed reactions are affected by heat denaturation of the enzyme protein molecule (2). This results in a dependence with an optimum. Not all enzymes are affected by temperature identically. This can be used for the separation of some specific enzymes. The activity of an enzyme mixture is estimated at a lower temperature, that of the more stable component is estimated at a raised temperature.

ACTIVATION

Of- E N Z Y M E S

1. By cleavage of an oligopeptide from a proenzyme. 2. By formation of SS bonds, resulting in the exposure of the active site (activation of ribonuclease by subtilisin). 3. By formation of a complex with metal ions. 4. By allosteric activation.

E N Z Y M E ACTIVATION AND FORMATION OFTHE ACTIVE SITE

The case of chymotrypsin is used to demonstrate these concepts. Under the action of pepsin or autocatalytically, two dipeptides are split from the originally single polypeptide chain with 249 amino acids of chymotrypsinogen (Ser14-Arg15 and Thr147--Asn148) which gives rise to three peptide chains interconnected by five SS bridges. This necessarily changes the conformation of the molecule and an active site of the enzyme is formed. Chymotrypsin has thus been formed from chymotrypsinogenThe active site is formed by Ser1'95, His87 and Asp1"2. A similar molecule, including the amino acid sequence of some parts of the protein chain, is that of trypsinogen (239 amino acids). It is activated to trypsin by splitting off a hexapeptide from the amino end of the chain. The conformation of the trypsin molecule is stabilized by six SS bonds which link together a single polypeptide chain.

ALLOSTERICENZYMES

are oligomers formed by two or more monomers. They can exist in two extreme, reversible states, called active and inactive. Every ligand (substrate, effector) that can form a complex with the protein can bind to each of the protein subunits: to an active site for the substrate, and to a regulatory site for the effector. Binding of the effector to a subunit is followed by a gradual change of conformation of other subunits. This results in a gradual transition of the allosteric enzyme to the active state. If the active state is changed back to the inactive one, the binding site for substrate changes its activity. As the protein passes from one conformation to the other, its molecular symmetry is preserved. Allosteric enzymes participate in the control of metabolism in multi-enzyme systems. The control of a metabolic sequence is effected by a negative or positive feedback.

1. States of an allosteric enzyme caused by a change of spatial organization. 2. Allosteric activation through binding an activator (positive effector). 3. Allosteric inhibition due to binding an inhibitor (negative effector).

MECHANISM OF E N Z Y M E ACTION
Mechanism of acetylcholine cleavage by cho-linesterase The active site of the enzyme contains two functionally and spatially separated sites:

1. COO- where the positively charged N+ of the substrate is bound electrostatically j 2. the active site proper possessing an esterase activity, with Ser, , His.
The substrate acetylcholine is bound in the reaction in a definite position in the_ active site by an ionic bond between the negative charge of COO-of the active site and the positively charged quaternary nitrogen N+= of the substrate. During the reaction, a proton dissociated from the phenol group of (active site) combines with the oxygen of the originally alcoholic group of choline. This generates a positive charge at the carbon of the substrate acetyl group which is attracted to the negatively charged oxygen of the dissociated alcoholic group of serine in the active site. The bond between (choline) and (acetyl) is broken and the liberated acetyl reacts with serine to a transient acetylserine. The proton split off from serine is attracted to the negatively charged oxygen of the dissociated phenol group of tyrosine, a hydroxy group is formed and the original state of the tyrosine residue of the active site is restored. The hydrolysis proper begins by dissociation of a water molecule, the H+ being attracted to the nitrogen of the imidazole group of His, the hydroxyl is set free and attacks the transiently formed ester bond of acetyl-serine. This liberates a molecule of acetic acid. The H+ bound transiently to the imidazole nitrogen is set free and reattached to the O- of serine. The original state of all the three active-site amino acids is restored. The released choline and acetic acid leave the active centre by diffusion. All the processes described above take place more or less simultaneously. The hydrolysis of acetylcholine results from a combined action of all the functional groups of the active site. The reaction course is affected by the distances between the individual subsites in the active centre. Model experiments done with substrate analogs indicate that the reaction will proceed smoothly if the basic group of imidazole lies 0.5 am from the COO- group of acetyl, and if the tyrosine chain lies 0.25 nm from the side chain of serine. This provides further evidence for the importance of the active site conformation for its activity.

K I N E T I C S OF E N Z Y M E CATALYZED R E A C T I O N S At a given enzyme concentration the reaction rate depends on the concentration of substrate. This substrate dependence of the rate is graphically described by a rectangular hyperbola such that at low concentrations of substrate the reaction is first-order while at very high substrate concentrations it is zero-order. This observation was the basis for the formulation of a fundamental theory of enzyme kinetics, by L. Michaelis and M. L. Menten in 1913. It is based on the assumption that an enzyme-substrate complex is formed which undergoes a chemical reaction and breaks down to the free snzyme and a product (eq. 1). A characteristic value of every enzyme reaction, the , can be derived as follows. The actual enzyme concentration after the complex has been formed is [E] = [E(] [ES]. The overall reaction rate depends on the rates at which the ES complex is formed and broken down. The re-formation of the ES complex from the enzyme and the product is usually neglected. At steady state, the rate of ES formation vi and the rate of its decomposition 2 are identical and thus expression 2 may be used. The rate of product formation is given by v &a[ES]. The value of [ES] can be computed from eq. 3 and substituted in eq. 5. The maximum rate Kmax is attained at high substrate concentrations when all the enzyme molecules are saturated. Then [ES] = [Ej] and eq. 6 can be used. Eq. 5 can now be rearranged by USmg max- The KM can be estimated from measurable quantities, substrate concentration, reaction rate, and maximum reaction rate (eq. 7). If v = * , eq. 8 can be applied, showing that has the dimension of concentration. The KM. value depends on the type of substrate, the pH of the reaction mixture and the temperature. If an enzyme catalyzes the reaction of several related substrates, every substrate will have its characteristic KM- In an approximation, reactions with a lower KM will proceed more readily. ' L I N E A R I Z A T I O N Of THE M I C H A E L S EQUATiOK

(Lineweaver and Burk)

The reaction rate v is expressed by eq 9. If a reciprocal of the equation is taken, it has the form of (10) and, after rearrangement, of (11) which is an equation of the straight line = a + bx where = 1/v; a = 1/Kmax (this determines the intercept with the y-axis); b = / (this is the slope of the straight line); 1/[S].

COMPETITIVE

INHIBITION

is reversible and can be relieved by a high concentration of substrate. It is made possible by a lower specificity of the binding site of the enzyme so that structurally related compounds can be bound but not necessarily chemically transformed. The reaction rate depends on the ratio of inhibitor and substrate concentrations and on their relative affinity for the enzyme. The apparent value increases.

NONCOMPETITIVE

INHIBITION

cannot be relieved by excess substrate. The inhibitor binds allotopically with respect to substrate and is not structurally related to the substrate. The reaction rate depends on the concentration of inhibitor and on the inhibition constant Ki. The apparent Fmai decreases as (1 + [l]lKi) rises.

ALLOSTERIC

INHIBITION

is either reversible or irreversible. It is caused by a negative effector (inhibitor) being bound at a site different from the active site (a regulatory site). The inhibitor is often the final reaction product of a multi-enzyme system. The dependence of the reaction rate on the concentrations of substrate and inhibitor is not simple and cannot be expressed as a change of - This is connected with the fact that the dependence of initial rate on substrate concentration in allosteric enzymes usually has an Sshaped character.

Clan 1. Oxidoreductases

Subgroup

Reaction catalyzed hydrogenation and dehydrogenation

ENZYME CLASSIFICATION

1.1 1.2 1.3 1.4 1.5 1.6

When coining names of enzymes, the substrate name was usually taken and the suffix -ase was attached (arginase catalyzes the hydrolysis of arginine; phosphatase hydro lyzes phosphoric esters, etc.). In other cases, the suffix was attached to the name of the catalyzed reaction, thus: dehydrogenase catalyzes dehydrogenations, hydrolase catalyzes hydrolysis, transferase a transfer of a chemical group, etc. Some of the early described enzymes have special names, such as trypsin, pepsin, catalase. As the number of enzymes known keeps increasing, the International Union of Biochemistry recommended to introduce a decimal system of enzymes based on the nature of the catalyzed reaction. Such a classification provides direct information on the character of the catalyzed process. In 1972, the Commission for Biochemical Nomenclature of the International Union of Pure and Applied Chemistry (IUPAC) published a new edition of Enzyme Nomenclature (1973). The first number in the classification describe the main class (2 stands for Transferases), the next number refers to some characteristic of the reaction (2.1 indicates the transfer of a one-carbon residue), the next number provides further details (2.1.1 means transfer of a methyl group). Both systematic and trivial (but logically constructed) names are now in use.

2. Transferases 2.1 2.2 2.3 2.4 2.5 2.6 2.7 2.8 3. Hydrolases 3.1 3.2 3.3 3.4 3.5 3.6 4. Lyases 4.1 4.2 4.3 5. Isomerases 5.1 5.2 5.3 5.4 6. Ligases 6.1 6.2 6.3 6.4

transfer of functional groups one-carbon residues aldehyde or keto group acyl glycosyl bond nonmethyl alkyl or aryl nitrogen-containing group phosphorus-containing groups sulfur-cntaining groups

hydrolytic reactions of esters glycosides ethers peptides other C-N bonds acid anhydrides addition to a double bond

isomerizations racemases and epimerases cistraru isomerases intramolecular oxidoreductases intromalecular transferases bond formation using ATP

OXIDOREDUCTASES Several types of enzyme-catalyzed oxidationreduction reactions. The enzymes transfer hydrogen or electrons and catalyze biological oxidations. They contain specific coenzymes. They are grouped according to the donor from which they accept hydrogen or electron, or according to the acceptor to which they transmit it.

Enzyme

Group transferred

General reaction

TRANSFERASES transfer groups of atoms by specific carriers which act as coenzymes. They play a role in biochemical conversions and can transfer methyl, carboxyl, amino, sulfo, formyl (Ci) or phosphoryl groups.

Phosphor ransf erase Phosphoryl H1PO4

Aminotransferase

Amino NHj

Sulfotransferase

Sulfuryl -SO3H

(transferase

Acetyl, succinyl. aminoacyl

Enzyme Peptidases

Substrate Proteins Peptides

Bond attached Peptide

General reaction

Glycoside hydrolases

Polysaccharides disaccharides

Glycoside

Esterases Lipases Phosphodiesterases

Neutral lipids phospholipids Polynucleotide

Ester

Phosphodlester

HYDROLASES
Phosphatase s Phosphate esters Ph os ph noes te*

catalyze' hydrolytic cleavage and are named according to the bond they attack (glycosidases, esterases, etc.).

Enzyme Decarboxylase

Group removed CO,

General reaction

LYASES are enzymes splitting groups from the substrate molecule nonhydrolytically; also they form double bonds or assist in the addition of groups to double bonds. They can split off carbon dioxide, water, ammonia, and larger groups.

Aldol&se

Lyase or synthase for Ketoicid reverie reaction

Dehydratase

HiO

Deaminase

NH3

Enzyme

Group isomerteed Carbonyl

Alternative position of group C-1 --------* C-2

General reaction

ISOMERASES

Glucose 6-phosphate fsomerase

catalyze interconversions of isomers. This may include racemization, cis-trans isomerization, shift of a double bond, exchange of groups on an asymmetric carbon, shift of a phosphate group to another carbon, etc.

Phosphoglycerate phospho mutase

Phosphoryl

C-2 <*- C-3

Race mase

Hydroxyl and hydrogen

D ---------* L

Enzyme Aminoacyl-tRNA synthetase

Bond formed C-0

Example Amino acid activation protein synthesis

Glutamine synthetase

C-N

Glutamine biosynthesis

Amino acid ligase

C-N

Protein synthesis

Li G A S E S

Acetyl CoA carboxylase

C-C

Synthesis of fatty acids (formation of malonyl CoA)

are enzymes synthesizing bonds with the aid of the high energy PP bond. Reaction partners may be ATP or another high-energy compound, or biotin during carboxylation reactions.