AChE bivalent inhibitors

From Proteopedia

Revision as of 13:50, 25 January 2009

PART I

This page is a continuation of the page "AChE inhibitors and substrates".

The of TcAChE consists of two binding subsites. First of them is the "catalytic anionic site" (CAS), which involves mentioned above catalytic triad (colored orange) and the conserved residues and also participating in ligands recognition. Another conserved residue (colored cyan) is situated at the second binding subsite, termed the "peripheral anionic site" (PAS), ~14 Å from CAS. Therefore, the ligands that will be able to interact with both these subsites, will be more potent AChE inhibitors in comparison to compounds interacting only with CAS. One of the ways to produce such ligands is to introduce two active substances into one compound. If it is spatially necessary these subunits could be divided by alkyl linker with suitable length. According to the strategy of the use of a bivalent ligand, the (RS)-(±)-tacrine-(10)-hupyridone ((R)-3 or (S)-3) was designed and synthesized. It consists of mentioned in the page 'AChE inhibitors and substrates' (colored magenta), 10-carbon (yellow), and (red). The tacrine moiety of this inhibitor binds at the CAS, the linker spans the gorge, and the hupyridone moiety binds at the PAS.

The comparison of the (R)-3/TcAChE and tacrine/TcAChE complexes at the . A of the trigonal (R)-3/TcAChE structure (1zgb; (R)-3 colored cyan; TcAChE residues interacting with (R)-3 are colored sea-green) with the crystal structure of tacrine/TcAChE (1acj, (tacrine colored magenta; residues interacting with tacrine are colored pink) reveals a similar binding mode for the tacrine moiety. In both structures the tacrine ring is situated at the CAS, between the aromatic residues Trp84 and Phe330. Steric clash with the 10-carbon linker could explain the tilt observed for the Phe330 (yellow and transparent in the tacrine/TcAChE). Water molecules are shown as red spheres. The tacrine unit of (R)-3 N forms with His440 O (3.0 Å) similar to that of tacrine alone. Similarly to the tacrine/TcAChE structure the system of three water molecules at the CAS ((R)-3/TcAChE) binds the tacrine-linker N via hydrogen bonds to Ser81 O, Ser122 Oγ, and Asn85 Oδ1 (2.6-3.5 Å) .

The comparison of the (R)-3/TcAChE (1zgb) and bis-hupyridone/TcAChE complexes (1h22 and 1h23) at the . of the (R)-tacrine-(10)-hupyridone ((R)-3, cyan) and (S,S)-(-)-Bis(12)-hupyridone ((S,S)-(-)-4b, orange, i.e. 12-carbon-tether-linked hupyridone dimer) and (S,S)-(-)-Bis(10)-hupyridone ((S,S)-(-)-4a, plum) complexes demonstrates the binding mode of the hupyridone moiety. TcAChE residues of symmetry-related molecule are shown in magenta. X-ray structures of TcAChE complexed with these 10- and 12-carbon-tether-linked 2 (S,S)-(-)-4a and (S,S)-(-)-4b show one subunit bound at the , the linker spanning the gorge, and the other subunit bound at the . There are two connecting the hupyridone O to Lys11 Nζ and hupyridone N to Gln185 Oε1 of a symmetry-related molecule at (R)-3/TcAChE complex. Water molecules are shown as red spheres. Another hydrogen bond connects the hupyridone O to a water molecule, which is bound to Ser286 N. Similarly, the hupyridone-PAS unit of both (-)-4a and (-)-4b forms direct and an indirect hydrogen bonds with the protein backbone in the PAS region.

The of (R)-3 (cyan) and (S)-3 (1zgc) bound to the TcAChE active site in the orthorhombic forms is shown. The residues important for inhibitor binding are in green. In contrast to the trigonal form (1zgb), the residues Gln*185 and Lys*11 (yellow) of an other symmetry-related monomer do not form hydrogen bonds with the ligands. of (S,S)-(-)-4a (1h22) and (S)-3, orthorhombic TcAChE (1zgc) demonstrates the similar mode of binding of the hupyridone unit at the PAS. The residues Trp279 (top) and Trp84 (bottom) represent the PAS and the CAS, respectively.

Described in the page 'AChE inhibitors and substrates' (GAL; colored red) is an AChE inhibitor and it is currently used in therapy of the AD. Conjugate of GAL through (8 carbons, yellow) with a (blueviolet) called compound 3 has a larger affinity for AChE than that of GAL alone. This is similar to previously described cases of bivalent ligands.

A comparison between the in complex with TcAChE and the structure (1dx6) shows an identical binding mode of the GAL-moiety (transparent red) of compound 3 to that of GAL alone (blue) at the CAS. The PEG molecule (gray) is located at the active site of galanthamine/TcAChE structure. The alkyl linker spans the active-site gorge and phthalimido moiety of the compound 3 is situated near the Trp279 at the PAS. The compound 3 has larger affinity to TcAChE than GAL. It could be explained by increased number of interactions between compound 3 (which interacts not only with residues within CAS but also within PAS) and TcAChE in comparison to GAL.

The (yellow) interacted with 13 residues of the the from Trp84 at the bottom to Phe284 at the top. Nine of these residues were (Tyr70, Trp84, Tyr121, Trp279, Phe284, Phe330, Phe331, Tyr334, and His440; colored darkmagenta). The contacts made by the drug in the bottom of the gorge involved with Trp84, Tyr121, Phe331, and His440 and, especially, a stacking interaction with Phe330. The carbamate moiety of CPT-11 was located near the residues . So, the carbon C9 (shown in magenta) of the carbamate linkage in CPT-11, was 9.3 Å from Oγ, the nucleophilic atom within the three catalytic residues Ser200, His440, and Glu327. The steric clashes between CPT-11 and TcAChE residues prevented positioning CPT-11 near the Ser200 Oγ (where hydrolysis could occur), therefore, TcAChE can not hydrolyze CPT-11.

Among the most interesting drugs that have been designed to inhibit acetylcholinesterase are those that have two binding sites that bind both the peripheral and catatylic sites simultaneously. Such drugs bind highly specificly and strongly. A good example is .

It appears that the principal interaction between the aceylcholine and the enzyme is relatively newly discovered cation-pi interactions between the cationic moiety of the substrate and the many aromatic residues lining the catalytic gorge. Unlike most interatomic interactions in chemistry, cation-pi interactions are unusual in that their energy hardly changes as the cationic and aromatic ring centers vary between 4 and 7 Angstroms apart, and for a wide variety of relative orientations of the aromatic rings. This gives the substrate an energetically smooth ride down the gorge with few bumps or barriers to impede passage down the gorge.

Most acetylcholinesterases have a net negative charge and a large patch of negative potential around the entrance to the active site gorge, which may be useful to attract the positively charged acetycholine substrate to the site. As one travels down the gorge, this potential becomes increasingly more and more negative, reaching a peak at the active site at the base. Because of this potential, the peripherial site is thought to act like a substrate trap, that forces practically molecule of substrate that reaches the peripheral site to travel down the gorge to the active site, that probably contributes greatly to the extremely rapid rate of degrading the substrate. This whole enzyme therefore acts like a brilliantly designed natural vacuum cleaner that clears the neurotransmitter out of the synapse extremely quickly. Yet to be solved, however, is how the products clear the active site rapidly, whether back through the gorge, or out a back door on the other side of the protein that quickly opens each catalytic cycle (Trp 84 is actually near the surface of the 'underside' of the protein.)

For information about additional AChE inhibitors see page AChE bivalent inhibitors (Part II).

References

Atomic structure of acetylcholinesterase from Torpedo californica: a prototypic acetylcholine-binding protein., Sussman JL, Harel M, Frolow F, Oefner C, Goldman A, Toker L, Silman I, Science. 1991 Aug 23;253(5022):872-9. PMID:1678899

The complex of a bivalent derivative of galanthamine with torpedo acetylcholinesterase displays drastic deformation of the active-site gorge: implications for structure-based drug design., Greenblatt HM, Guillou C, Guenard D, Argaman A, Botti S, Badet B, Thal C, Silman I, Sussman JL, J Am Chem Soc. 2004 Dec 1;126(47):15405-11. PMID:15563167

The crystal structure of the complex of the anticancer prodrug 7-ethyl-10-[4-(1-piperidino)-1-piperidino]-carbonyloxycamptothecin (CPT-11) with Torpedo californica acetylcholinesterase provides a molecular explanation for its cholinergic action., Harel M, Hyatt JL, Brumshtein B, Morton CL, Yoon KJ, Wadkins RM, Silman I, Sussman JL, Potter PM, Mol Pharmacol. 2005 Jun;67(6):1874-81. Epub 2005 Mar 16. PMID:15772291