Transition state analog

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Transition state analogs (transition state analogues), are chemical compounds with a chemical structure that resembles the transition state of a substrate molecule in an enzyme-catalyzed chemical reaction. Enzymes interact with a substrate by means of strain or distortions, moving the substrate towards the transition state. [1] Transition state analogs can be used as inhibitors in enzyme-catalyzed reactions by blocking the active site of the enzyme. Theory suggests that enzyme inhibitors which resembled the transition state structure would bind more tightly to the enzyme than the actual substrate. [2] Examples of drugs that are transition state analog inhibitors include flu medications such as the neuraminidase inhibitor oseltamivir and the HIV protease inhibitors saquinavir in the treatment of AIDS.

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Transition state analogue

Enzyme-catalyzed reactions lower the overall activation energy of a reaction Enzyme action.png
Enzyme-catalyzed reactions lower the overall activation energy of a reaction

The transition state of a structure can best be described in regards to statistical mechanics where the energies of bonds breaking and forming have an equal probability of moving from the transition state backwards to the reactants or forward to the products. In enzyme-catalyzed reactions, the overall activation energy of the reaction is lowered when an enzyme stabilizes a high energy transition state intermediate. Transition state analogs mimic this high energy intermediate but do not undergo a catalyzed chemical reaction and can therefore bind much stronger to an enzyme than simple substrate or product analogs.

Designing transition state analogue

To design a transition state analogue, the pivotal step is the determination of transition state structure of substrate on the specific enzyme of interest with experimental method, for example, kinetic isotope effect. In addition, the transition state structure can also be predicted with computational approaches as a complementary to KIE. We will explain these two methods in brief.

Kinetic isotope effect

Kinetic isotope effect (KIE) is a measurement of the reaction rate of isotope-labeled reactants against the more common natural substrate. Kinetic isotope effect values are a ratio of the turnover number and include all steps of the reaction. [3] Intrinsic kinetic isotope values stem from the difference in the bond vibrational environment of an atom in the reactants at ground state to the environment of the atom's transition state. [3] Through the kinetic isotope effect much insight can be gained as to what the transition state looks like of an enzyme-catalyzed reaction and guide the development of transition state analogs.

Computational simulation

Computational approaches have been regarded as a useful tool to elucidate the mechanism of action of enzymes. [4] Molecular mechanics itself can not predict the electron transfer which is the fundamental of organic reaction but the molecular dynamics simulation provide sufficient information considering the flexibility of protein during catalytic reaction. The complementary method would be combined molecular mechanics/ quantum mechanics simulation (QM/MM)methods. [5] With this approach, only the atoms responsible for enzymatic reaction in the catalytic region will be reared with quantum mechanics and the rest of the atoms were treated with molecular mechanics. [6]

Examples of transition state analogue design

After determining the transition state structures using either KIE or computation simulations, the inhibitor can be designed according to the determined transition state structures or intermediates. The following three examples illustrate how the inhibitors mimic the transition state structure by changing functional groups correspond to the geometry and electrostatic distribution of the transition state structures.

Methylthioadenosine nucleosidase inhibitor

Transition state analogue example one TSA 1.png
Transition state analogue example one

Methylthioadenosine nucleosidase are enzymes that catalyse the hydrolytic deadenylation reaction of 5'-methylthioadenosine and S-adenosylhomocysteine. It is also regarded as an important target for antibacterial drug discovery because it is important in the metabolic system of bacteria and only produced by bacteria. [7] Given the different distance between nitrogen atom of adenine and the ribose anomeric carbon (see in the diagram in this section), the transition state structure can be defined by early or late dissociation stage. Based on the finding of different transition state structures, Schramm and coworkers designed two transition state analogues mimicking the early and late dissociative transition state. The early and late transition state analogue shown binding affinity (Kd) of 360 and 140 pM, respectively. [8]

Thermolysin inhibitor

Transitions state analogue example 2 TS3.png
Transitions state analogue example 2

Thermolysin is an enzyme produced by Bacillus thermoproteolyticus that catalyses the hydrolysis of peptides containing hydrophobic amino acids. [9] Therefore, it is also a target for antibacterial agents. The enzymatic reaction mechanism starts form the small peptide molecule and replaces the zinc binding water molecule towards Glu143 of thermolysin. The water molecule is then activated by both the zinc ion and the Glu143 residue and attacks the carbonyl carbon to form a tetrahedral transition state (see figure). Holden and coworkers then mimicked that tetrahedral transition state to design a series of phosphonamidate peptide analogues. Among the synthesized analogues, R = L-Leu possesses the most potent inhibitory activity (Ki = 9.1 nM). [10]

Arginase inhibitor

Transition state analogue example 3 TSA3.png
Transition state analogue example 3

Arginase is a binuclear manganese metalloprotein that catalyses the hydrolysis of L-arginine to L-ornithine and urea. It is also regarded as a drug target for the treatment of asthma. [11] The mechanism of hydrolysis of L-arginine is carried out via nucleophilic attack on the guanidino group by water, forming a tetrahedral intermediate. Studies shown that a boronic acid moiety adopts a tetrahedral configuration and serves as an inhibitor. In addition, the sulfonamide functional group can also mimic the transition state structure. [12] Evidence of boronic acid mimics as transition state analogue inhibitors of human arginase I was elucidated by x-ray crystal structures. [13]

See also

Related Research Articles

Enzyme Large biological molecule that acts as a catalyst

Enzymes are proteins that act as biological catalysts (biocatalysts). Catalysts accelerate chemical reactions. The molecules upon which enzymes may act are called substrates, and the enzyme converts the substrates into different molecules known as products. Almost all metabolic processes in the cell need enzyme catalysis in order to occur at rates fast enough to sustain life. Metabolic pathways depend upon enzymes to catalyze individual steps. The study of enzymes is called enzymology and a new field of pseudoenzyme analysis has recently grown up, recognising that during evolution, some enzymes have lost the ability to carry out biological catalysis, which is often reflected in their amino acid sequences and unusual 'pseudocatalytic' properties.

Active site Active region of an enzyme

In biology, 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 and residues that catalyse a reaction of that substrate. Although the active site occupies only ~10–20% of the volume of an enzyme, it is the most important part as it directly catalyzes the chemical reaction. It usually consists of three to four amino acids, while other amino acids within the protein are required to maintain the tertiary structure of the enzymes.

Arginase

Arginase (EC 3.5.3.1, arginine amidinase, canavanase, L-arginase, arginine transamidinase) is a manganese-containing enzyme. The reaction catalyzed by this enzyme is: arginine + H2O → ornithine + urea. It is the final enzyme of the urea cycle. It is ubiquitous to all domains of life.

In physical organic chemistry, a kinetic isotope effect (KIE) is the change in the reaction rate of a chemical reaction when one of the atoms in the reactants is replaced by one of its isotopes. Formally, it is the ratio of rate constants for the reactions involving the light (kL) and the heavy (kH) isotopically substituted reactants (isotopologues):

A tetrahedral intermediate is a reaction intermediate in which the bond arrangement around an initially double-bonded carbon atom has been transformed from trigonal to tetrahedral. Tetrahedral intermediates result from nucleophilic addition to a carbonyl group. The stability of tetrahedral intermediate depends on the ability of the groups attached to the new tetrahedral carbon atom to leave with the negative charge. Tetrahedral intermediates are very significant in organic syntheses and biological systems as a key intermediate in esterification, transesterification, ester hydrolysis, formation and hydrolysis of amides and peptides, hydride reductions, and other chemical reactions.

Enzyme kinetics Study of biochemical reaction rates catalysed by an enzyme

Enzyme kinetics is the study of the rates of enzyme-catalysed chemical reactions. In enzyme kinetics, the reaction rate is measured and the effects of varying the conditions of the reaction are investigated. Studying an enzyme's kinetics in this way can reveal the catalytic mechanism of this enzyme, its role in metabolism, how its activity is controlled, and how a drug or a modifier might affect the rate.

Frank Westheimer

Frank Henry Westheimer was an American chemist. He taught at the University of Chicago from 1936 to 1954, and at Harvard University from 1953 to 1983, becoming the Morris Loeb Professor of Chemistry in 1960, and Professor Emeritus in 1983. The Westheimer medal was established in his honor in 2002.

Enzyme inhibitor Molecule that binds to an enzyme and decreases its activity

An enzyme inhibitor is a molecule that binds to an enzyme and decreases its activity. By binding to enzymes' active sites, inhibitors reduce the compatibility of substrate and enzyme and this leads to the inhibition of Enzyme-Substrate complexes' formation, preventing the catalysis of reactions and decreasing the amount of product produced by a reaction. It can be said that as the concentration of enzyme inhibitors increases, the rate of enzyme activity decreases, and thus, the amount of product produced is inversely proportional to the concentration of inhibitor molecules. Since blocking an enzyme's activity can kill a pathogen or correct a metabolic imbalance, many drugs are enzyme inhibitors. They are also used in pesticides. Not all molecules that bind to enzymes are inhibitors; enzyme activators bind to enzymes and increase their enzymatic activity, while enzyme substrates bind and are converted to products in the normal catalytic cycle of the enzyme.

Phosphoglycerate kinase

Phosphoglycerate kinase is an enzyme that catalyzes the reversible transfer of a phosphate group from 1,3-bisphosphoglycerate (1,3-BPG) to ADP producing 3-phosphoglycerate (3-PG) and ATP :

Enzyme catalysis Catalysis of chemical reactions by specialized proteins known as enzymes

Enzyme catalysis is the increase in the rate of a process by a biological molecule, an "enzyme". Most enzymes are proteins, and most such processes are chemical reactions. Within the enzyme, generally catalysis occurs at a localized site, called the active site.

Transition state theory

Transition state theory (TST) explains the reaction rates of elementary chemical reactions. The theory assumes a special type of chemical equilibrium (quasi-equilibrium) between reactants and activated transition state complexes.

Carboxypeptidase A

Carboxypeptidase A usually refers to the pancreatic exopeptidase that hydrolyzes peptide bonds of C-terminal residues with aromatic or aliphatic side-chains. Most scientists in the field now refer to this enzyme as CPA1, and to a related pancreatic carboxypeptidase as CPA2.

Chorismate mutase

In enzymology, chorismate mutase is an enzyme that catalyzes the chemical reaction for the conversion of chorismate to prephenate in the pathway to the production of phenylalanine and tyrosine, also known as the shikimate pathway. Hence, this enzyme has one substrate, chorismate, and one product, prephenate. Chorismate mutase is found at a branch point in the pathway. The enzyme channels the substrate, chorismate to the biosynthesis of tyrosine and phenylalanine and away from tryptophan. Its role in maintaining the balance of these aromatic amino acids in the cell is vital. This is the single known example of a naturally occurring enzyme catalyzing a pericyclic reaction. Chorismate mutase is only found in fungi, bacteria, and higher plants. Some varieties of this protein may use the morpheein model of allosteric regulation.

D-lysine 5,6-aminomutase

In enzymology, D-lysine 5,6-aminomutase is an enzyme that catalyzes the chemical reaction

In enzymology, an adenosylhomocysteine nucleosidase (EC 3.2.2.9) is an enzyme that catalyzes the chemical reaction

In enzymology, a methylthioadenosine nucleosidase (EC 3.2.2.16) is an enzyme that catalyzes the chemical reaction

Substrate analogs, are chemical compounds with a chemical structure that resemble the substrate molecule in an enzyme-catalyzed chemical reaction. Substrate analogs can act as competitive inhibitors of an enzymatic reaction. An example is phosphoramidate to the Tetrahymena group I ribozyme

Supramolecular catalysis Field of chemistry

Supramolecular catalysis is not a well-defined field but it generally refers to an application of supramolecular chemistry, especially molecular recognition and guest binding, toward catalysis. This field was originally inspired by enzymatic system which, unlike classical organic chemistry reactions, utilizes non-covalent interactions such as hydrogen bonding, cation-pi interaction, and hydrophobic forces to dramatically accelerate rate of reaction and/or allow highly selective reactions to occur. Because enzymes are structurally complex and difficult to modify, supramolecular catalysts offer a simpler model for studying factors involved in catalytic efficiency of the enzyme. Another goal that motivates this field is the development of efficient and practical catalysts that may or may not have an enzyme equivalent in nature.

Vaborbactam

Vaborbactam (INN) is a non-β-lactam β-lactamase inhibitor discovered by Rempex Pharmaceuticals, a subsidiary of The Medicines Company. While not effective as an antibiotic by itself, it restores potency to existing antibiotics by inhibiting the β-lactamase enzymes that would otherwise degrade them. When combined with an appropriate antibiotic it can be used for the treatment of gram-negative bacterial infections.

Position-specific isotope analysis

Position-specific isotope analysis, also called site-specific isotope analysis, is a branch of isotope analysis aimed at determining the isotopic composition of a particular atom position in a molecule. Isotopes are elemental variants with different numbers of neutrons in their nuclei, thereby having different atomic masses. Isotopes are found in varying natural abundances depending on the element; their abundances in specific compounds can vary from random distributions due to environmental conditions that act on the mass variations differently. These differences in abundances are called "fractionations," which are characterized via stable isotope analysis.

References

  1. ↑ Silverman, Richard B. (2004). The Organic Chemistry of Drug Design and Drug Action . San Diego, CA: Elsevier Academic Press. ISBN   0-12-643732-7.
  2. ↑ Copeland, R.A.; Davis, J.P.; Cain, G.A.; Pitts, W.J.; Magolda, R.L. (1996). "The Immunosuppressive Metabolite of Leflunomide is a Potent Inhibitor of Human Dihydroorotate Dehydrogenase". Biochemistry. 35 (4): 1270–3. doi:10.1021/bi952168g. PMID   8573583.
  3. 1 2 Schramm, Vern L (2011). "Enzymatic Transition States, Transition-State Analogs, Dynamics, Thermodynamics, and Lifetimes". Annu. Rev. Biochem. 80 (1): 703–732. doi:10.1146/annurev-biochem-061809-100742. PMC   5502542 . PMID   21675920.
  4. ↑ Peter, Kollman; Kuhn, B.; Peräkylä, M. (2002). "Computational Studies of Enzyme-Catalyzed Reactions: Where Are We in Predicting Mechanisms and in Understanding the Nature of Enzyme Catalysis?". J. Phys. Chem. B. 106 (7): 1537–1542. doi:10.1021/jp012017p.
  5. ↑ Hou, G; Hou, G.; Cui, Q. (2011). "QM/MM Analysis Suggests that Alkaline Phosphatase (AP) and Nucleotide Pyrophosphatase/Phosphodiesterase Slightly Tighten the Transition State for Phosphate Diester Hydrolysis Relative to Solution: Implication for Catalytic Promiscuity in the AP Superfamily". J. Am. Chem. Soc. 134 (1): 229–246. doi:10.1021/ja205226d. PMC   3257412 . PMID   22097879.
  6. ↑ Schwartz, S; Saen-oon, S.; Quaytman-Machleder, S.; Schramm, V. L.; Schwartz, S. D. (2008). "Atomic Detail of Chemical Transformation at the Transition State of an Enzymatic Reaction". PNAS. 105 (43): 16543–16545. Bibcode:2008PNAS..10516543S. doi: 10.1073/pnas.0808413105 . PMC   2575456 . PMID   18946041.
  7. ↑ Singh, Vipender; Singh V; Lee JE; Núñez S; Howell PL; Schramm VL. (2005). "Transition state structure of 5'-methylthioadenosine/S-adenosylhomocysteine nucleosidase from Escherichia coli and its similarity to transition state analogues". Biochemistry. 44 (35): 11647–11659. doi:10.1021/bi050863a. PMID   16128565.
  8. ↑ Guitierrez, Jemy; Luo, M.; Singh, V.; Li, L.; Brown, R. L.; Norris, G. E. (2007). "Picomolar Inhibitors as Transition-State Probes of 5′-Methylthioadenosine Nucleosidases". ACS Chemical Biology. 2 (11): 725–734. doi:10.1021/cb700166z. PMID   18030989.
  9. ↑ S, Endo (1962). "Studies on protease produced by thermophilic bacteria". J. Ferment. Technol. 40: 346–353.
  10. ↑ Holden, Hazel; Tronrud, D. E.; Monzingo, A. F.; Weaver, L. H. (1987). "Slow-and fast-binding inhibitors of thermolysin display different modes of binding: crystallographic analysis of extended phosphoramidate transition-state analogs". Biochemistry. 26 (26): 8542–8553. doi:10.1021/bi00400a008. PMID   3442675.
  11. ↑ Maarsingh, Harm; Johan Zaagsma; Herman Meurs (October 2009). "Arginase: a key enzyme in the pathophysiology of allergic asthma opening novel therapeutic perspectives". Br J Pharmacol. 158 (3): 652–664. doi:10.1111/j.1476-5381.2009.00374.x. PMC   2765587 . PMID   19703164.
  12. ↑ E, Cama; Shin H; Christianson DW. (2003). "Design of amino acid sulfonamides as transition-state analogue inhibitors of arginase". J Am Chem Soc. 125 (43): 13052–7. doi:10.1021/ja036365b. PMID   14570477.
  13. ↑ Shishova, Ekaterina; Luigi Di Costanzo; David E. Cane; David W. Christianson (2009). "Probing the Specificity Determinants of Amino Acid Recognition by Arginase". Biochemistry. 48 (1): 121–131. doi:10.1021/bi801911v. PMC   2665027 . PMID   19093830.
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