Inhibitors、抑制劑

Definition

Inhibitors are molecules that interact in some way with the enzyme to prevent it from working in the normal manner. There are a variety of types of inhibitors including: nonspecific, irreversible, reversible - competitive and noncompetitive.

Discovery

In 1965, Umezawa H initiated the screening study of enzyme inhibitors produced by microorganisms and isolated leupeptin and antipain inhibiting trypsin and papain, chymostatin inhibiting chymotrypsin, pepstatin inhibiting pepsin, panosialin inhibiting sialidases, oudenone inhibiting tyrosine hydroxylase, dopastin inhibiting dopamine 3-hydroxylase, aquayamycin and chrothiomycin inhibiting tyrosine hydroxylase and dopamine J3-hydroxylase 1. Recently, an alternative approach has been applied to predict new inhibitors: rational drug design uses the three-dimensional structure of an enzyme's active site to predict which molecules might be inhibitors 2.

Structural Characteristics

The crystal structures of many inhibitors have been determined. The crystal structures of three highly potent and selective low-molecular weight rigid peptidyl aldehyde inhibitors complexed with thrombin have been determined. All the three inhibitors have a novel lactam moiety at the P3 position, while the two with greatest trypsin selectivity have a guanidinopiperidyl group at the P1 position that binds in the S1 specificity site. The kinetics of inhibition vary from slow to fast with thrombin and are fast in all cases with trypsin. The kinetics are examined in terms of the slow formation of a stable transition-state complex in a two-step mechanism 3. Protein tyrosine phosphatases PTPN5, PTPRR and PTPN7 comprise a family of phosphatases that specifically inactivate MAPKs (mitogen-activated protein kinases). Structures of these enzymes along with two classes of inhibitors, cyclopenta[c]quinolinecarboxylic acids and 2,5-dimethylpyrrolyl benzoic acids have been determined. One of the identified inhibitor scaffolds, cyclopenta[c]quinoline, was docked successfully into PTPRR. The structures together with the established structure-activity relationship propose new avenues for the development of selective inhibitors that have therapeutic potential for treating neurodegenerative diseases in the case of PTPRR or acute myeloblastic leukemia targeting PTPN7 4. N-terminal pyroglutamate (pGlu) formation from its glutaminyl (or glutamyl) precursor is required in the maturation of numerous bioactive peptides. The structure of human QC in free form and bound to a substrate and three imidazole-derived inhibitors reveals an alpha/beta scaffold akin to that of two-zinc exopeptidases but with several insertions and deletions, particularly in the active-site region. The relatively closed active site displays alternate conformations due to the different indole orientations of Trp-207, resulting in two substrate (glutamine t-butyl ester)-binding modes. The structural analyses of several active-site-mutant enzymes provide a structural basis for the rational design of inhibitors against QC-associated disorders 5.

Mode of Action

Inhibitor binding can stop a substrate from entering the enzyme's active site and/or hinder the enzyme from catalyzing its reaction. There are a variety of types of inhibitors including: nonspecific, irreversible, reversible - competitive and noncompetitive. Reversible inhibitors bind to enzymes with non-covalent interactions like hydrophobic interactions, hydrogen bonds, and ionic bonds. Multiple weak bonds between the inhibitor and the active site lead to strong and specific binding. Non-specific methods of inhibition include any physical or chemical changes which ultimately denature the protein portion of the enzyme and are therefore irreversible. Specific Inhibitors exert their effects upon a single enzyme. Most poisons work by specific inhibition of enzymes. A competitive inhibitor is any compound which closely resembles the chemical structure and molecular geometry of the substrate. The inhibitor competes for the same active site as the substrate molecule. The inhibitor may interact with the enzyme at the active site, but no reaction takes place. A noncompetitive inhibitor is a substance that interacts with the enzyme, but usually not at the active site.  The net effect of a non competitive inhibitor is to change the shape of the enzyme and thus the active site, so that the substrate can no longer interact with the enzyme to give a reaction. Non competitive inhibitors are usually reversible. Irreversible Inhibitors form strong covalent bonds with an enzyme.  These inhibitors may act at, near, or remote from the active site.  Consequently, they may not be displaced by the addition of excess substrate.

Functions

Drug molecules, many drug molecules are enzyme inhibitors and a medicinal enzyme inhibitor is usually characterized by its specificity and its potency. A high specificity and potency suggests that a drug will have fewer side effects and less toxic. Enzyme inhibitors are found in nature and are also designed and produced as part of pharmacology and biochemistry 6. Natural poisons are often enzyme inhibitors that have evolved to defend a plant or animal against predators. These natural toxins include some of the most poisonous compounds known. Artificial inhibitors are often used as drugs, but can also be insecticides such as malathion, herbicides such as glyphosate, or disinfectants such as triclosan.

Nerve gases such as diisopropylfluorophosphate (DFP) inhibit the active site of acetylcholine esterase by reacting with the hydroxyl group of serine to make an ester.

Inhibition of blood clotting, Oxalic and citric acid inhibit blood clotting by forming complexes with calcium ions necessary for the enzyme metal ion activator.

References

1.Umezawa. H (1972). Chemistry of enzyme inhibitors of microbial origin. University Park Press, Baltimore, Md. 21202.,114.

2.Scapin G (2006). Structural biology and drug discovery. Curr. Pharm. Des., 12 17): 2087–2097.

3. Krishnan R, Zhang E, Hakansson K, Arni RK, Tulinsky A, Lim-Wilby MS, Levy OE, Semple JE, Brunck TK (1998). Highly Selective Mechanism-Based Thrombin Inhibitors:  Structures of Thrombin and Trypsin Inhibited with Rigid Peptidyl Aldehydes. Biochemistry, 37(35):12094–12103.

4.Eswaran J , von Kries JP, Marsden B, Longman E, Debreczeni JE, Ugochukwu E, Turnbull A, Lee WH, Knapp S, Barr AJ (2006). Crystal structures and inhibitor identification for PTPN5, PTPRR and PTPN7: a family of human MAPK-specific protein tyrosine phosphatases. Biochem J., 395(3):483-491.

5.Huang KF, Liu YL, Cheng WJ, Ko TP, Wang AH (2005). Crystal structures of human glutaminyl cyclase, an enzyme responsible for protein N-terminal pyroglutamate formation. PNAS., 102(37):13117-13122.

6.Holmes CF, Maynes JT, Perreault KR, Dawson JF, James MN Curr Med Chem., 9(22):1981-1989.