Ariel N

General

Full name : Ariel Naomi

First name : Naomi

Mail : Israel Institute for Biological Research, PO Box 19, Ness-Ziona 70450

Zip Code :

City :

Country : Israel

Email : ariel@iibr.gov.il

Phone : 972-8-938-1529

Fax : 972-8-940-1404

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References (37)

Title : Surprising findings from the functional analysis of human acetylcholinesterase adducts of Alzheimer?s disease drugs. -
Author(s) : Ordentlich A , Barak D , Ariel N , Kronman C , Kaplan D , Velan B , Shafferman A
Ref : Cholinergic Mechanisms, CRC Press :177 , 2004
PubMedID:

Title : Attempts to engineer an enzyme mimic of butyrylcholinesterase by substitution of the six divergent aromatic amino acids in the active center of acetylcholinesterase. -
Author(s) : Kaplan D , Ordentlich A , Barak D , Ariel N , Kronman C , Baruch V , Shafferman A
Ref : Cholinergic Mechanisms, CRC Press :601 , 2004
PubMedID:

Title : Poster (14) The aromatic trapping of histidine 447 in catalysis of acetylcholinesterases -
Author(s) : Shafferman A , Barak D , Kaplan D , Ordentlich A , Ariel N , Velan B
Ref : In: Cholinesterases in the Second Millennium: Biomolecular and Pathological Aspects , (Inestrosa NC, Campos EO) P. Universidad Catolica de Chile-FONDAP Biomedicina :327 , 2004
PubMedID:

Title : The aromatic trapping of histidine 447 in catalysis of acetylcholinesterases -
Author(s) : Shafferman A , Barak D , Kaplan D , Ordentlich A , Ariel N , Velan B
Ref : In: Cholinesterases in the Second Millennium: Biomolecular and Pathological Aspects , (Inestrosa NC, Campos EO) P. Universidad Catolica de Chile-FONDAP Biomedicina :181 , 2004
PubMedID:

Title : Poster (21) Analysis of acetylcholinesterase adducts of alzheimer's drugs -
Author(s) : Ordentlich A , Kronman C , Barak D , Ariel N , Kaplan D , Velan B , Shafferman A
Ref : In: Cholinesterases in the Second Millennium: Biomolecular and Pathological Aspects , (Inestrosa NC, Campos EO) P. Universidad Catolica de Chile-FONDAP Biomedicina :332 , 2004
PubMedID:

Title : The aromatic trapping of the catalytic histidine is essential for efficient catalysis in acetylcholinesterase - Barak_2002_Biochemistry_41_8245
Author(s) : Barak D , Kaplan D , Ordentlich A , Ariel N , Velan B , Shafferman A
Ref : Biochemistry , 41 :8245 , 2002
Abstract : While substitution of the aromatic residues (Phe295, Phe338), located in the vicinity of the catalytic His447 in human acetylcholinesterase (HuAChE) had little effect on catalytic activity, simultaneous replacement of both residues by aliphatic amino acids resulted in a 680-fold decrease in catalytic activity. Molecular simulations suggested that the activity decline is related to conformational destabilization of His447, similar to that observed for the hexamutant HuAChE which mimics the active center of butyrylcholinesterase. On the basis of model structures of other cholinesterases (ChEs), we predicted that catalytically nonproductive mobility of His447 could be restricted by introduction of aromatic residue in a different location adjacent to this histidine (Val407). Indeed, the F295A/F338A/V407F enzyme is 170-fold more reactive than the corresponding double mutant and only 3-fold less reactive than the wild-type HuAChE. However, analogous substitution of Val407 in the hexamutant HuAChE (generating the heptamutant Y72N/Y124Q/W286A/F295L/F297V/Y337A/V407F) did not enhance catalytic activity. Reactivity of these double, triple, hexa, and hepta mutant HuAChEs was monitored toward covalent ligands such as organophosphates and the transition state analogue TMFTA, which probe, respectively, the facility of the enzymes to accommodate Michaelis complexes and to undergo the acylation process. The findings suggest that in the F295A/F338A mutant the two His447 conformational states, which are essential for the different stages of the catalytic process, seem to be destabilized. On the other hand, in the F295A/F338A/V407F mutant only the state involved in acylation is impaired. Such differential effects on the His447 conformational properties demonstrate the general role of aromatic residues in cholinesterases, and probably in other serine hydrolases, in "trapping" of the catalytic histidine and thereby in optimization of catalytic activity.
ESTHER : Barak_2002_Biochemistry_41_8245
PubMedSearch : Barak_2002_Biochemistry_41_8245
PubMedID: 12081473

Title : Does butyrylization of acetylcholinesterase through substitution of the six divergent aromatic amino acids in the active center gorge generate an enzyme mimic of butyrylcholinesterase? - Kaplan_2001_Biochemistry_40_7433
Author(s) : Kaplan D , Ordentlich A , Barak D , Ariel N , Kronman C , Velan B , Shafferman A
Ref : Biochemistry , 40 :7433 , 2001
Abstract : The active center gorge of human acetylcholinesterase (HuAChE) is lined by 14 aromatic residues, whereas in the closely related human butyrylcholinesterase (HuBChE) 3 of the aromatic active center residues (Phe295, Phe297, Tyr337) as well as 3 of the residues at the gorge entrance (Tyr72, Tyr124, Trp286) are replaced by aliphatic amino acids. To investigate whether this structural variability can account for the reactivity differences between the two enzymes, gradual replacement of up to all of the 6 aromatic residues in HuAChE by the corresponding residues in HuBChE was carried out. The affinities of the hexamutant (Y72N/Y124Q/W286A/F295L/F297V/Y337A) toward tacrine, decamethonium, edrophonium, huperzine A, or BW284C51 differed by about 5-, 80-, 170-, 25000-, and 17000-fold, respectively, from those of the wild-type HuAChE. For most of these prototypical noncovalent active center and peripheral site ligands, the hexamutant HuAChE displayed a reactivity phenotype closely resembling that of HuBChE. These results support the accepted view that the active center architectures of AChE and BChE differ mainly by the presence of a larger void space in BChE. Nevertheless, reactivity of the hexamutant HuAChE toward the substrates acetylthiocholine and butyrylthiocholine, or covalent ligands such as phosphonates and the transition state analogue m-(N,N,N-trimethylammonio)trifluoroacetophenone (TMTFA), is about 45-170-fold lower than that of HuBChE. Most of this reduction in reactivity can be related to the combined replacements of the three aromatic residues at the active center, Phe295, Phe297, and Tyr337. We propose that the hexamutant HuAChE, unlike BChE, is impaired in its capacity to accommodate certain tetrahedral species in the active center. This impairment may be related to the enhanced mobility of the catalytic histidine His447, which is observed in molecular dynamics simulations of the hexamutant and the F295L/F297V/Y337A HuAChE enzymes but not in the wild-type HuAChE.
ESTHER : Kaplan_2001_Biochemistry_40_7433
PubMedSearch : Kaplan_2001_Biochemistry_40_7433
PubMedID: 11412096

Title : Structures of recombinant native and E202Q mutant human acetylcholinesterase complexed with the snake-venom toxin fasciculin-II - Kryger_2000_Acta.Crystallogr.D.Biol.Crystallogr_56_1385
Author(s) : Kryger G , Harel M , Giles K , Toker L , Velan B , Lazar A , Kronman C , Barak D , Ariel N , Shafferman A , Silman I , Sussman JL
Ref : Acta Crystallographica D Biol Crystallogr , 56 :1385 , 2000
Abstract : Structures of recombinant wild-type human acetylcholinesterase and of its E202Q mutant as complexes with fasciculin-II, a 'three-finger' polypeptide toxin purified from the venom of the eastern green mamba (Dendroaspis angusticeps), are reported. The structure of the complex of the wild-type enzyme was solved to 2.8 A resolution by molecular replacement starting from the structure of the complex of Torpedo californica acetylcholinesterase with fasciculin-II and verified by starting from a similar complex with mouse acetylcholinesterase. The overall structure is surprisingly similar to that of the T. californica enzyme with fasciculin-II and, as expected, to that of the mouse acetylcholinesterase complex. The structure of the E202Q mutant complex was refined starting from the corresponding wild-type human acetylcholinesterase structure, using the 2.7 A resolution data set collected. Comparison of the two structures shows that removal of the charged group from the protein core and its substitution by a neutral isosteric moiety does not disrupt the functional architecture of the active centre. One of the elements of this architecture is thought to be a hydrogen-bond network including residues Glu202, Glu450, Tyr133 and two bridging molecules of water, which is conserved in other vertebrate acetylcholinesterases as well as in the human enzyme. The present findings are consistent with the notion that the main role of this network is the proper positioning of the Glu202 carboxylate relative to the catalytic triad, thus defining its functional role in the interaction of acetylcholinesterase with substrates and inhibitors.
ESTHER : Kryger_2000_Acta.Crystallogr.D.Biol.Crystallogr_56_1385
PubMedSearch : Kryger_2000_Acta.Crystallogr.D.Biol.Crystallogr_56_1385
PubMedID: 11053835
Gene_locus related to this paper: human-ACHE

Title : Exploring the active center of human acetylcholinesterase with stereomers of an organophosphorus inhibitor with two chiral centers - Ordentlich_1999_Biochemistry_38_3055
Author(s) : Ordentlich A , Barak D , Kronman C , Benschop HP , De Jong LP , Ariel N , Barak R , Segall Y , Velan B , Shafferman A
Ref : Biochemistry , 38 :3055 , 1999
Abstract : The stereoselectivity of the phosphonylation reaction and the effects of adduct configuration on the aging process were examined for human acetylcholinesterase (HuAChE) and its selected active center mutants, using the four stereomers of 1,2,2-trimethylpropyl methylphosphonofluoridate (soman). The reactivity of wild type HuAChE toward the PS-soman diastereomers was 4.0-7.5 x 10(4)-fold higher than that toward the PR-diastereomers. Aging of the PSCS-somanyl-HuAChE conjugate was also >1.6 x 10(4)-fold faster than that of the corresponding PRCS-somanyl adduct, as shown by both reactivation and electrospray mass spectrometry (ESI/MS) experiments. On the other hand, both processes exhibited very limited sensitivity to the chirality of the alkoxy group Calpha of either PS- or PR-diastereomers. These stereoselectivities presumably reflect the relative participation of the enzyme in stabilization of the Michaelis complexes and in dealkylation of the respective covalent conjugates, and therefore could be utilized for further probing of the HuAChE active center functional architecture. Reactivities of HuAChE enzymes carrying replacements at the acyl pocket (F295A, F297A, and F295L/F297V) indicate that stereoselectivity with respect to the soman phosphorus chirality depends on the structure of this binding subsite, but this stereoselectivity cannot be explained only by limitation in the capacity to accommodate the PR-diastereomers. In addition, these acyl pocket enzyme mutants display some (5-10-fold) preference for the PRCR-soman over the PRCS-stereomer, while reactivity of the hydrophobic pocket mutant enzyme W86F toward the PRCS-soman resembles that of the wild type HuAChE. Residue substitutions in the H-bond network (E202Q, E450A, Y133F, and Y133A) and the hydrophobic pocket (F338A, W86A, W86F, and Y337A) result in a limited stereoselectivity for the PSCS- over the PSCR-stereomer. Aging of the PS-somanyl conjugates with all the HuAChE mutant enzymes tested practically lacked stereoselectivity with respect to the Calpha of the alkoxy moiety. Thus, the inherent asymmetry of the active center does not seem to affect the rate-determining step of the dealkylation process, possibly because both the PSCS- and the PSCR-somanyl moieties yield the same carbocationic intermediate.
ESTHER : Ordentlich_1999_Biochemistry_38_3055
PubMedSearch : Ordentlich_1999_Biochemistry_38_3055
PubMedID: 10074358

Title : 3D Structure at 2.7 Resolution of Native and E202Q Mutant Human Acetylcholinesterase Complexed with Fasciculin-II -
Author(s) : Kryger G , Giles K , Harel M , Toker L , Velan B , Lazar A , Kronman C , Barak D , Ariel N , Shafferman A , Silman I , Sussman JL
Ref : In: Structure and Function of Cholinesterases and Related Proteins - Proceedings of Sixth International Meeting on Cholinesterases , (Doctor, B.P., Taylor, P., Quinn, D.M., Rotundo, R.L., Gentry, M.K. Eds) Plenum Publishing Corp. :323 , 1998
PubMedID:

Title : 3D Structure of a Complex of Human Acetylcholinesterase with Fasciculin-II at 2.7 Resolution -
Author(s) : Kryger G , Giles K , Harel M , Toker L , Velan B , Lazar A , Kronman C , Barak D , Ariel N , Shafferman A , Silman I , Sussman JL
Ref : In: Structure and Function of Cholinesterases and Related Proteins - Proceedings of Sixth International Meeting on Cholinesterases , (Doctor, B.P., Taylor, P., Quinn, D.M., Rotundo, R.L., Gentry, M.K. Eds) Plenum Publishing Corp. :370 , 1998
PubMedID:

Title : Compatibility of Structures Inferred from Mutagenesis and from X-Ray Crystallography for Various AChE Complexes -
Author(s) : Ariel N , Ordentlich A , Barak D , Bino T , Velan B , Shafferman A
Ref : In: Structure and Function of Cholinesterases and Related Proteins - Proceedings of Sixth International Meeting on Cholinesterases , (Doctor, B.P., Taylor, P., Quinn, D.M., Rotundo, R.L., Gentry, M.K. Eds) Plenum Publishing Corp. :375 , 1998
PubMedID:

Title : The 'aromatic patch' of three proximal residues in the human acetylcholinesterase active centre allows for versatile interaction modes with inhibitors - Ariel_1998_Biochem.J_335_95
Author(s) : Ariel N , Ordentlich A , Barak D , Bino T , Velan B , Shafferman A
Ref : Biochemical Journal , 335 :95 , 1998
Abstract : The role of the functional architecture of the human acetylcholinesterase (HuAChE) active centre in accommodating the non-covalent inhibitors tacrine and huperzine A, or the carbamates pyridostigmine and physostigmine, was analysed using 16 mutants of residues lining the active-centre gorge. Despite the structural diversity of the ligands, certain common properties of the complexes could be observed: (a) replacement of aromatic residues Tyr133, Tyr337 and especially Trp86, resulted in pronounced changes in stability of all the complexes examined; (b) effects due to replacements of the five other aromatic residues along the active-centre gorge, such as the acyl pocket (Phe295, Phe297) or at the peripheral anionic site (Tyr124, Trp286, Tyr341) were relatively small; (c) effects due to substitution of the carboxylic residues in the gorge (Glu202, Glu450) were moderate. These results and molecular modelling indicate that the aromatic side chains of residues Trp86, Tyr133 and Tyr337 form together a continuous 'aromatic patch' lining the wall of the active-centre gorge, allowing for the accommodation of the different ligands via multiple modes of interaction. Studies with HuAChE mutants carrying replacements at positions 86, 133 and 337 indicate that the orientations of huperzine A and tacrine in the HuAChE complexes in solution are significantly different from those observed in X-ray structures of the corresponding complexes with Torpedo californica AChE (TcAChE). These discrepancies may be explained in terms of structural differences between the complexes of HuAChE and TcAChE or, more likely, by the enhanced flexibility of the AChE active-centre gorge in solution as compared with the crystalline state.
ESTHER : Ariel_1998_Biochem.J_335_95
PubMedSearch : Ariel_1998_Biochem.J_335_95
PubMedID: 9742217

Title : Structural Modifications of the Omega Loop in Human Acetylcholinesterase -
Author(s) : Ariel N , Velan B , Barak D , Leitner M , Bino T , Ordentlich A , Shafferman A
Ref : In: Structure and Function of Cholinesterases and Related Proteins - Proceedings of Sixth International Meeting on Cholinesterases , (Doctor, B.P., Taylor, P., Quinn, D.M., Rotundo, R.L., Gentry, M.K. Eds) Plenum Publishing Corp. :453 , 1998
PubMedID:

Title : Functional characteristics of the oxyanion hole in human acetylcholinesterase - Ordentlich_1998_J.Biol.Chem_273_19509
Author(s) : Ordentlich A , Barak D , Kronman C , Ariel N , Segall Y , Velan B , Shafferman A
Ref : Journal of Biological Chemistry , 273 :19509 , 1998
Abstract : The contribution of the oxyanion hole to the functional architecture and to the hydrolytic efficiency of human acetylcholinesterase (HuAChE) was investigated through single replacements of its elements, residues Gly-121, Gly-122 and the adjacent residue Gly-120, by alanine. All three substitutions resulted in about 100-fold decrease of the bimolecular rate constants for hydrolysis of acetylthiocholine; however, whereas replacements of Gly-120 and Gly-121 affected only the turnover number, mutation of residue Gly-122 had an effect also on the Michaelis constant. The differential behavior of the G121A and G122A enzymes was manifested also toward the transition state analog m-(N,N, N-trimethylammonio)trifluoroacetophenone (TMTFA), organophosphorous inhibitors, carbamates, and toward selected noncovalent active center ligands. Reactivity of both mutants toward TMTFA was 2000-11, 000-fold lower than that of the wild type HuAChE; however, the G121A enzyme exhibited a rapid inhibition pattern, as opposed to the slow binding kinetics shown by the G122A enzyme. For both phosphates (diethyl phosphorofluoridate, diisopropyl phosphorofluoridate, and paraoxon) and phosphonates (sarin and soman), the decrease in inhibitory activity toward the G121A enzyme was very substantial (2000-6700-fold), irrespective of size of the alkoxy substituents on the phosphorus atom. On the other hand, for the G122A HuAChE the relative decline in reactivity toward phosphonates (500-460-fold) differed from that toward the phosphates (12-95-fold). Although formation of Michaelis complexes with substrates does not seem to involve significant interaction with the oxyanion hole, interactions with this motif are a major stabilizing element in accommodation of covalent inhibitors like organophosphates or carbamates. These observations and molecular modeling suggest that replacements of residues Gly-120 or Gly-121 by alanine alter the structure of the oxyanion hole motif, abolishing the H-bonding capacity of residue at position 121. These mutations weaken the interaction between HuAChE and the various ligands by 2.7-5.0 kcal/mol. In contrast, variations in reactivity due to replacement of residue Gly 122 seem to result from steric hindrance at the active center acyl pocket
ESTHER : Ordentlich_1998_J.Biol.Chem_273_19509
PubMedSearch : Ordentlich_1998_J.Biol.Chem_273_19509
PubMedID: 9677373

Title : Contribution of the Active Center Functional Architecture to AChE Reactivity Toward Substrates and Inhibitors -
Author(s) : Shafferman A , Ordentlich A , Barak D , Kronman C , Ariel N , Velan B
Ref : In: Structure and Function of Cholinesterases and Related Proteins - Proceedings of Sixth International Meeting on Cholinesterases , (Doctor, B.P., Taylor, P., Quinn, D.M., Rotundo, R.L., Gentry, M.K. Eds) Plenum Publishing Corp. :203 , 1998
PubMedID:

Title : Does Electrostatic Attraction or Steering by Charged Residues within the Gorge Contribute to the Reactivity of AChE? -
Author(s) : Ordentlich A , Barak D , Stein D , Berman D , Kronman C , Ariel N , Velan B , Shafferman A
Ref : In: Structure and Function of Cholinesterases and Related Proteins - Proceedings of Sixth International Meeting on Cholinesterases , (Doctor, B.P., Taylor, P., Quinn, D.M., Rotundo, R.L., Gentry, M.K. Eds) Plenum Publishing Corp. :234 , 1998
PubMedID:

Title : Bovine acetylcholinesterase: cloning, expression and characterization - Mendelson_1998_Biochem.J_334_251
Author(s) : Mendelson I , Kronman C , Ariel N , Shafferman A , Velan B
Ref : Biochemical Journal , 334 :251 , 1998
Abstract : The bovine acetylcholinesterase (BoAChE) gene was cloned from genomic DNA and its structure was determined. Five exons coding for the AChE T-subunit and the alternative H-subunit were identified and their organization suggests high conservation of structure in mammalian AChE genes. The deduced amino acid sequence of the bovine T-subunit is highly similar to the human sequence, showing differences at 34 positions only. However, the cloned BoAChE sequence differs from the published amino acid sequence of AChE isolated from fetal bovine serum (FBS) by: (1) 13 amino acids, 12 of which are conserved between BoAChE and human AChE, and (2) the presence of four rather than five potential N-glycosylation sites. The full coding sequence of the mature BoAChE T-subunit was expressed in human embryonal kidney 293 cells (HEK-293). The catalytic properties of recombinant BoAChE and its reactivity towards various inhibitors were similar to those of the native bovine enzyme. Soluble recombinant BoAChE is composed of monomers, dimers and tetramers, yet in contrast to FBS-AChE, tetramer formation is not efficient. Comparative SDS/PAGE analysis reveals that all four potential N-glycosylation sites identified by DNA sequencing appear to be utilized, and that recombinant BoAChE comigrates with FBS-AChE. A major difference between the recombinant enzyme and the native enzyme was observed when clearance from circulation was examined. The HEK-293-derived enzyme was cleared from the circulation at a much faster rate than FBS-AChE. This difference in behaviour, together with previous studies on the effect of post-translation modification on human AChE clearance [Kronman, Velan, Marcus, Ordentlich, Reuveny and Shafferman (1995) Biochem. J. 311, 959-967] suggests that cell-dependent glycosylation plays a key role in AChE circulatory residence.
ESTHER : Mendelson_1998_Biochem.J_334_251
PubMedSearch : Mendelson_1998_Biochem.J_334_251
PubMedID: 9693127
Gene_locus related to this paper: bovin-ACHE

Title : Aging of somanyl-acetylcholinesterase adducts: facts and models [letter] -
Author(s) : Shafferman A , Ordentlich A , Barak D , Stein D , Ariel N , Velan B
Ref : Biochemical Journal , 324 :996 , 1997
PubMedID: 9235881

Title : Structural modifications of the omega loop in human acetylcholinesterase - Velan_1996_FEBS.Lett_395_22
Author(s) : Velan B , Barak D , Ariel N , Leitner M , Bino T , Ordentlich A , Shafferman A
Ref : FEBS Letters , 395 :22 , 1996
Abstract : Conformational mobility of the surface omega loop (Cys-69-Cys-96) in human acetylcholinesterase (HuAChE) was recently implicated in substrate accessibility to the active center and in the mechanism of allosteric modulation of enzymatic activity. We therefore generated and kinetically evaluated the following modifications or replacements in HuAChE: (a) residues at the loop ends, (b) residues involved in putative hydrogen-bond interactions within the loop and between the loop and the protein core, (c) ChEs conserved proline residues within the loop and (d) a deletion of a conserved segment of 5 residues. All the residue replacements, including those of the prolines, had either limited or no effect on enzyme reactivity. These results suggest that unlike the case of lipase, the omega loop in the HuAChE is not involved in large lid-like displacements. In cases where modifications of the loop sequence had some effect on reactivity, the effects could be attributed to an altered position of residue Trp-86 supporting the proposed coupling between the structure of the omega loop and the positioning of the Trp-86 indole moiety, in catalytic activity and in allosterism.
ESTHER : Velan_1996_FEBS.Lett_395_22
PubMedSearch : Velan_1996_FEBS.Lett_395_22
PubMedID: 8849682

Title : Aging of phosphylated human acetylcholinesterase: catalytic processes mediated by aromatic and polar residues of the active centre - Shafferman_1996_Biochem.J_318_833
Author(s) : Shafferman A , Ordentlich A , Barak D , Stein D , Ariel N , Velan B
Ref : Biochemical Journal , 318 :833 , 1996
Abstract : We have examined the effects of 11 substitutions of active centre gorge residues of human acetylcholinesterase (HuAChE) on the rates of phosphonylation by 1,2,2-trimethylpropyl methyl-phosphonofluoridate (soman) and the aging of the resulting conjugates. The rates of phosphonylation were reduced to as little as one-seventieth, mainly in mutants of the hydrogen-bond network (Glu-202, Glu-450, Tyr-133). These recombinant enzymes as well as the F338A, W86A, W86F and D74N mutant HuAChEs varied in their resistance to aging (15-3300-fold relative to the wild type). The most dramatic resistance to aging was observed for the phosphonyl conjugate of the mutant W86A enzyme (1850-3300-fold relative to the wild type). It is proposed that Trp-86 contributes to the aging process by stabilizing the evolving carbonium ion on the 1,2,2-trimethylpropyl moiety, via charge-pi interaction. The rate-enhancing effect of Trp-86 provides a rationale for the unique facility of aging in soman-inhibited cholinesterases, compared with the corresponding conjugates in other serine hydrolases. Replacements of Glu-202 by aspartic acid, glutamine or alanine residues resulted in a similar (1/130-1/300) decrease of the rates of aging. A comparable decrease was also observed for the conjugate of the F338A mutant. These results, and the similar pH dependence of aging rates for the wild-type and E202Q and F338A mutant HuAChEs, indicate that Glu-202 is not involved in proton transfer to the phosphonyl moiety. On the basis of these findings and of molecular modelling we suggest that Glu-202 and Phe-338 contribute to the aging process by stabilizing the imidazolium of the catalytic triad His-447 via charge-charge and charge-pi interactions respectively, thereby facilitating an oxonium formation on the phosphonyl moiety.
ESTHER : Shafferman_1996_Biochem.J_318_833
PubMedSearch : Shafferman_1996_Biochem.J_318_833
PubMedID: 8836126

Title : The architecture of human acetylcholinesterase active center probed by interactions with selected organophosphate inhibitors - Ordentlich_1996_J.Biol.Chem_271_11953
Author(s) : Ordentlich A , Barak D , Kronman C , Ariel N , Segall Y , Velan B , Shafferman A
Ref : Journal of Biological Chemistry , 271 :11953 , 1996
Abstract : The role of the functional architecture of human acetylcholinesterase (HuAChE) active center in facilitating reactions with organophosphorus inhibitors was examined by a combination of site-directed mutagenesis and kinetic studies of phosphorylation with organophosphates differing in size of their alkoxy substituents and in the nature of the leaving group. Replacements of residues Phe-295 and Phe-297, constituting the HuAChE acyl pocket, increase up to 80-fold the reactivity of the enzymes toward diisopropyl phosphorofluoridate, diethyl phosphorofluoridate, and p-nitrophenyl diethyl phosphate (paraoxon), indicating the role of this subsite in accommodating the phosphate alkoxy substituent. On the other hand, a decrease of up to 160-fold in reactivity was observed for enzymes carrying replacements of residues Tyr-133, Glu-202, and Glu-450, which are constituents of the hydrogen bond network in the HuAChE active center, which maintains its unique functional architecture. Replacement of residues Trp-86, Tyr-337, and Phe-338 in the alkoxy pocket affected reactivity toward diisopropyl phosphorofluoridate and paraoxon, but to a lesser extent that toward diethyl phosphorofluoridate, indicating that both the alkoxy substituent and the p-nitrophenoxy leaving group interact with this subsite. In all cases the effects on reactivity toward organophosphates, demonstrated in up to 10,000-fold differences in the values of bimolecular rate constants, were mainly a result of altered affinity of the HuAChE mutants, while the apparent first order rate constants of phosphorylation varied within a narrow range. This finding indicates that the main role of the functional architecture of HuAChE active center in phosphorylation is to facilitate the formation of enzyme-inhibitor Michaelis complexes and that this affinity, rather than the nucleophilic activity of the enzyme catalytic machinery, is a major determinant of HuAChE reactivity toward organophosphates.
ESTHER : Ordentlich_1996_J.Biol.Chem_271_11953
PubMedSearch : Ordentlich_1996_J.Biol.Chem_271_11953
PubMedID: 8662593

Title : Molecular Aspects of Catalysis and of Allosteric Regulation of Aceytlcholinesterases -
Author(s) : Shafferman A , Ordentlich A , Barak D , Kronman C , Ariel N , Leitner M , Segall Y , Bromberg A , Reuveny S , Marcus D , Bino T , Lazar A , Cohen S , Velan B
Ref : In Enzyme of the Cholinesterase Family - Proceedings of Fifth International Meeting on Cholinesterases , (Quinn, D.M., Balasubramanian, A.S., Doctor, B.P., Taylor, P., Eds) Plenum Publishing Corp. :189 , 1995
PubMedID:

Title : Contribution of aromatic moieties of tyrosine 133 and of the anionic subsite tryptophan 86 to catalytic efficiency and allosteric modulation of acetylcholinesterase - Ordentlich_1995_J.Biol.Chem_270_2082
Author(s) : Ordentlich A , Barak D , Kronman C , Ariel N , Segall Y , Velan B , Shafferman A
Ref : Journal of Biological Chemistry , 270 :2082 , 1995
Abstract : Substitution of Trp-86, in the active center of human acetylcholinesterase (HuAChE), by aliphatic but not by aromatic residues resulted in a several thousandfold decrease in reactivity toward charged substrate and inhibitors but only a severalfold decrease for noncharged substrate and inhibitors. The W86A and W86E HuAChE enzymes exhibit at least a 100-fold increase in the Michaelis-Menten constant or 100-10,000-fold increase in inhibition constants toward various charged inhibitors, as compared to W86F HuAChE or the wild type enzyme. On the other hand, replacement of Glu-202, the only acidic residue proximal to the catalytic site, by glutamine resulted in a nonselective decrease in reactivity toward charged and noncharged substrates or inhibitors. Thus, the quaternary nitrogen groups of substrates and other active center ligands, are stabilized by cation-aromatic interaction with Trp-86 rather than by ionic interactions, while noncharged ligands appear to bind to distinct site(s) in HuAChE. Analysis of the Y133F and Y133A HuAChE mutated enzymes suggests that the highly conserved Tyr-133 plays a dual role in the active center: (a) its hydroxyl appears to maintain the functional orientation of Glu-202 by hydrogen bonding and (b) its aromatic moiety maintains the functional orientation of the anionic subsite Trp-86. In the absence of aromatic interactions between Tyr-133 and Trp-86, the tryptophan acquires a conformation that obstructs the active site leading, in the Y133A enzyme, to several hundredfold decrease in rates of catalysis, phosphorylation, or in affinity to reversible active site inhibitors. It is proposed that allosteric modulation of acetylcholinesterase activity, induced by binding to the peripheral anionic sites, proceeds through such conformational change of Trp-86 from a functional anionic subsite state to one that restricts access of substrates to the active center.
ESTHER : Ordentlich_1995_J.Biol.Chem_270_2082
PubMedSearch : Ordentlich_1995_J.Biol.Chem_270_2082
PubMedID: 7836436

Title : Electrostatic Attraction by Surface Charge does not Contribute to the Catalytic Efficiency of Acetylcholinesterase -
Author(s) : Barak D , Ordentlich A , Kronman C , Ber R , Bino T , Ariel N , Osman R , Velan B , Shafferman A
Ref : In Enzyme of the Cholinesterase Family - Proceedings of Fifth International Meeting on Cholinesterases , (Quinn, D.M., Balasubramanian, A.S., Doctor, B.P., Taylor, P., Eds) Plenum Publishing Corp. :223 , 1995
PubMedID:

Title : Amino Acids Determining Specificity to OP-Agents and Facilitating the Aging Process in Human Acetylcholinesterase -
Author(s) : Ordentlich A , Kronman C , Stein D , Ariel N , Reuveny S , Marcus D , Segall Y , Barak D , Velan B , Shafferman A
Ref : In Enzyme of the Cholinesterase Family - Proceedings of Fifth International Meeting on Cholinesterases , (Quinn, D.M., Balasubramanian, A.S., Doctor, B.P., Taylor, P., Eds) Plenum Publishing Corp. :221 , 1995
PubMedID:

Title : Allosteric modulation of acetylcholinesterase activity by peripheral ligands involves a conformational transition of the anionic subsite - Barak_1995_Biochemistry_34_15444
Author(s) : Barak D , Ordentlich A , Bromberg A , Kronman C , Marcus D , Lazar A , Ariel N , Velan B , Shafferman A
Ref : Biochemistry , 34 :15444 , 1995
Abstract : Replacement of residues Asp74, Trp286, and Tyr72, which are constituents of the peripheral anionic site (PAS) of human acetylcholinesterase (HuAChE), affected similarly both the binding and the inhibition constants of the PAS-specific ligand propidium, demonstrating that changes in the inhibitory activity are a direct consequence of altered binding to the PAS. In contrast, the active center HuAChE mutants W86A and Y133A show respective 350- and 25-fold increased resistance to inhibition by propidium but no change in binding affinities, demonstrating that the allosteric mechanism of PAS-mediated inhibition involves a conformational change of these Trp86 and Tyr133 residues rather than physical obstruction of substrate access by the inhibitor itself. These findings support the recent proposal that the allosteric mechanism operates via transition between active and nonactive conformations of the anionic subsite Trp86 and that replacement of Tyr133 by alanine may stabilize a nonactive Trp86 conformation that occludes the active center [Ordentlich et al. (1995) J. Biol. Chem. 270, 2082]. In further support of this mechanism and the role of Tyr133, we find that (a) the dissociation constants (Kd) for the noncovalent complexes of the irreversible inhibitors diisopropyl phosphorofluoridate or paraoxon with Y133A HuAChE are increased 20-500-fold, relative to either wild-type enzyme or its Y133F or W86A mutants; and (b) access of substrates such as 3,3-dimethylbutyl thioacetate is restored by removal of Trp86 from the Y133A enzyme (i.e., the W86A/Y133A mutant). We suggest that the conformational transition of Trp86 is coupled to the motions of the cysteine loop (Cys69-Cys96) of HuAChE and is inherent to the dynamics of the native enzyme.
ESTHER : Barak_1995_Biochemistry_34_15444
PubMedSearch : Barak_1995_Biochemistry_34_15444
PubMedID: 7492545

Title : Acetylcholinesterase peripheral anionic site degeneracy conferred by amino acid arrays sharing a common core - Barak_1994_J.Biol.Chem_269_6296
Author(s) : Barak D , Kronman C , Ordentlich A , Ariel N , Bromberg A , Marcus D , Lazar A , Velan B , Shafferman A
Ref : Journal of Biological Chemistry , 269 :6296 , 1994
Abstract : Several of the residues constituting the peripheral anionic site (PAS) in human acetylcholinesterase (HuAChE) were identified by a combination of kinetic studies with 19 single and multiple HuAChE mutants, fluorescence binding studies with the Trp-286 mutant, and by molecular modeling. Mutants were analyzed with three structurally distinct positively charged PAS ligands, propidium, decamethonium, and di(p-allyl-N-dimethylaminophenyl)pentane-3-one (BW284C51), as well as with selective active center inhibitors, hexamethonium and edrophonium. Single mutations of residues Tyr-72, Tyr-124, Glu-285, Trp-286, and Tyr-341 resulted in up to 10-fold increase in inhibition constants for PAS ligands, whereas for multiple mutants up to 400-fold increase was observed. The 6th PAS element residue Asp-74 is unique in its ability to affect conformation of both the active site and the PAS (Shafferman, A., Velan, B., Ordentlich, A., Kronman, C., Grosfeld, H., Leitner, M., Flashner, Y., Cohen, S., Barak, D., and Ariel, N. (1992) EMBO J. 11, 3561-3568) as demonstrated by the several hundred-fold increase in Ki for D74N inhibition by the bisquaternary ligands decamethonium and BW284C51. Based on these studies, singular molecular models for the various HuAChE inhibitor complexes were defined. Yet, for the decamethonium complex two distinct conformations were generated, accommodating the quaternary ammonium group by interactions with either Trp-286 or with Tyr-341. We propose that the PAS consists of a number of binding sites, close to the entrance of the active site gorge, sharing residues Asp-74 and Trp-286 as a common core. Binding of ligands to these residues may be the key to the allosteric modulation of HuAChE catalytic activity. This functional degeneracy is a result of the ability of the Trp-286 indole moiety to interact either via stacking, aromatic-aromatic, or via pi-cation attractions and the involvement of the carboxylate of Asp-74 in charge-charge or H-bond interactions.
ESTHER : Barak_1994_J.Biol.Chem_269_6296
PubMedSearch : Barak_1994_J.Biol.Chem_269_6296
PubMedID: 8119978

Title : Electrostatic attraction by surface charge does not contribute to the catalytic efficiency of acetylcholinesterase - Shafferman_1994_EMBO.J_13_3448
Author(s) : Shafferman A , Ordentlich A , Barak D , Kronman C , Ber R , Bino T , Ariel N , Osman R , Velan B
Ref : EMBO Journal , 13 :3448 , 1994
Abstract : Acetylcholinesterases (AChEs) are characterized by a high net negative charge and by an uneven surface charge distribution, giving rise to a negative electrostatic potential extending over most of the molecular surface. To evaluate the contribution of these electrostatic properties to the catalytic efficiency, 20 single- and multiple-site mutants of human AChE were generated by replacing up to seven acidic residues, vicinal to the rim of the active-center gorge (Glu84, Glu285, Glu292, Asp349, Glu358, Glu389 and Asp390), by neutral amino acids. Progressive simulated replacement of these charged residues results in a gradual decrease of the negative electrostatic potential which is essentially eliminated by neutralizing six or seven charges. In marked contrast to the shrinking of the electrostatic potential, the corresponding mutations had no significant effect on the apparent bimolecular rate constants of hydrolysis for charged and non-charged substrates, or on the Ki value for a charged active center inhibitor. Moreover, the kcat values for all 20 mutants are essentially identical to that of the wild type enzyme, and the apparent bimolecular rate constants show a moderate dependence on the ionic strength, which is invariant for all the enzymes examined. These findings suggest that the surface electrostatic properties of AChE do not contribute to the catalytic rate, that this rate is probably not diffusion-controlled and that long-range electrostatic interactions play no role in stabilization of the transition states of the catalytic process.
ESTHER : Shafferman_1994_EMBO.J_13_3448
PubMedSearch : Shafferman_1994_EMBO.J_13_3448
PubMedID: 8062821

Title : Engineering resistance to 'aging' of phosphylated human acetylcholinesterase. Role of hydrogen bond network in the active center - Ordentlich_1993_FEBS.Lett_334_215
Author(s) : Ordentlich A , Kronman C , Barak D , Stein D , Ariel N , Marcus D , Velan B , Shafferman A
Ref : FEBS Letters , 334 :215 , 1993
Abstract : Recombinant human acetylcholinesterase (HuAChE) and selected mutants (E202Q, Y337A, E450A) were studied with respect to catalytic activity towards charged and noncharged substrates, phosphylation with organophosphorus (OP) inhibitors and subsequent aging of the OP-conjugates. Amino acid E450, unlike residues E202 and Y337, is not within interaction distance from the active center. Yet, the bimolecular rates of catalysis and phosphylation are 30-100 fold lower for both E450A and E202Q compared to Y337A or the wild type and in both mutants the resulting OP-conjugates show striking resistance to aging. It is proposed that a hydrogen bond network, that maintains the functional architecture of the active center, involving water molecules and residues E202 and E450, is responsible for the observed behaviour.
ESTHER : Ordentlich_1993_FEBS.Lett_334_215
PubMedSearch : Ordentlich_1993_FEBS.Lett_334_215
PubMedID: 8224249

Title : Direct observation and elucidation of the structures of aged and nonaged phosphorylated cholinesterases by 31P NMR spectroscopy - Segall_1993_Biochemistry_32_13441
Author(s) : Segall Y , Waysbort D , Barak D , Ariel N , Doctor BP , Grunwald J , Ashani Y
Ref : Biochemistry , 32 :13441 , 1993
Abstract : 31P NMR spectroscopy of butyrylcholinesterase (BChE), acetylcholinesterase (AChE), and chymotrypsin (Cht) inhibited by pinacolyl methylphosphonofluoridate (soman), methylphosphonodifluoridate (MPDF), and diisopropyl phosphorofluoridate (DFP) allowed direct observation of the OP-linked moiety of aged (nonreactivatable) and nonaged organophosphorus (OP)-ChE conjugates. The 31P NMR chemical shifts of OP-ChE conjugates clearly demonstrated insertion of a P-O- bond into the active site of aged OP-ChE adducts. The OP moiety of nonaged OP-ChEs was shown to be uncharged. The OP-bound pinacolyl moiety of soman-inhibited and aged AChE was detached completely, whereas only partial dealkylation of the pinacolyl group was observed for soman-inhibited BChEs. This suggests that the latter enzyme reacted with the less active stereoisomer(s) of soman. In the case of soman-inhibited Cht, no dealkylation could be experimentally detected for any of the four stereoisomers of OP-Cht adducts. Results are consistent with the contention that the phenomenon of enzyme-catalyzed dealkylation of OP adducts of serine hydrolases strongly depends on the orientation of both the catalytic His and the carboxyl side chain of either Glu or Asp positioned next to the catalytic Ser. The denatured protein of aged OP-ChE or OP-Cht is a convenient leaving group in nucleophilic displacements of tetrahedral OP compounds despite the presence of a P-O- bond. This indicates that the unusual resistance to reactivation of the aged enzyme cannot be ascribed to simple electrostatic repulsion of an approaching nucleophile. The broadening of the 31P NMR signal of native OP-ChEs relative to that of OP-Cht is in agreement with the crystal structure of AChE, showing that the active site region of ChEs in solution resides in a deep, narrow gorge.
ESTHER : Segall_1993_Biochemistry_32_13441
PubMedSearch : Segall_1993_Biochemistry_32_13441
PubMedID: 8257680

Title : Dissection of the human acetylcholinesterase active center determinants of substrate specificity. Identification of residues constituting the anionic site, the hydrophobic site, and the acyl pocket - Ordentlich_1993_J.Biol.Chem_268_17083
Author(s) : Ordentlich A , Barak D , Kronman C , Flashner Y , Leitner M , Segall Y , Ariel N , Cohen S , Velan B , Shafferman A
Ref : Journal of Biological Chemistry , 268 :17083 , 1993
Abstract : Substrate specificity determinants of human acetylcholinesterase (HuAChE) were identified by combination of molecular modeling and kinetic studies with enzymes mutated in residues Trp-86, Trp-286, Phe-295, Phe-297, Tyr-337, and Phe-338. The substitution of Trp-86 by alanine resulted in a 660-fold decrease in affinity for acetythiocholine but had no effect on affinity for the isosteric uncharged substrate (3,3-dimethylbutylthioacetate). The results demonstrate that residue Trp-86 is the anionic site which binds, through cation-pi interactions, the quaternary ammonium of choline, and that of active center inhibitors such as edrophonium. The results also suggest that in the non-covalent complex, charged and uncharged substrates with a common acyl moiety (acetyl) bind to different molecular environments. The hydrophobic site for the alcoholic portion of the covalent adduct (tetrahedral intermediate) includes residues Trp-86, Tyr-337, and Phe-338, which operate through nonpolar and/or stacking interactions, depending on the substrate. Substrates containing choline but differing in the acyl moiety (acetyl, propyl, and butyryl) revealed that residues Phe-295 and Phe-297 determine substrate specificity of the acyl pocket for the covalent adducts. Phe-295 also determines substrate specificity in the non-covalent enzyme substrate complex and thus, the HuAChE F295A mutant exhibits over 130-fold increase in the apparent bimolecular rate constant for butyrylthiocholine compared with wild type enzyme. Reactivity toward specific butyrylcholinesterase inhibitors is similarly dependent on the nature of residues at positions 295 and 297. Amino acid Trp-286 at the rim of the active site "gorge" and Trp-86, in the active center, are essential elements in the mechanism of inhibition by propidium, a peripheral anionic site ligand. Molecular modeling and kinetic data suggest that a cross-talk between Trp-286 and Trp-86 can result in reorientation of Trp-86 which may then interfere with stabilization of substrate enzyme complexes. It is proposed that the conformational flexibility of aromatic residues generates a plasticity in the active center that contributes to the high efficiency of AChE and its ability to respond to external stimuli.
ESTHER : Ordentlich_1993_J.Biol.Chem_268_17083
PubMedSearch : Ordentlich_1993_J.Biol.Chem_268_17083
PubMedID: 8349597
Gene_locus related to this paper: human-ACHE , human-BCHE

Title : Recombinant human acetylcholinesterase - Enzyme engineering -
Author(s) : Shafferman A , Velan B , Barak D , Kronman C , Ordentlich A , Flashner Y , Leitner M , Segal Y , Grosfeld H , Stein D , Ariel N
Ref : Medical Defense Bioscience Review , 3 :1097 , 1993
PubMedID:

Title : Mutagenesis of human acetylcholinesterase. Identification of residues involved in catalytic activity and in polypeptide folding - Shafferman_1992_J.Biol.Chem_267_17640
Author(s) : Shafferman A , Kronman C , Flashner Y , Leitner M , Grosfeld H , Ordentlich A , Gozes Y , Cohen S , Ariel N , Barak D , Harel M , Silman I , Sussman JL , Velan B
Ref : Journal of Biological Chemistry , 267 :17640 , 1992
Abstract : Evidence for the involvement of Ser-203, His-447, and Glu-334 in the catalytic triad of human acetylcholinesterase was provided by substitution of these amino acids by alanine residues. Of 20 amino acid positions mutated so far in human acetylcholinesterase (AChE), these three were unique in abolishing detectable enzymatic activity (less than 0.0003 of wild type), yet allowing proper production, folding, and secretion. This is the first biochemical evidence for the involvement of a glutamate in a hydrolase triad (Schrag, J.D., Li, Y., Wu, M., and Cygler, M. (1991) Nature 351, 761-764), supporting the x-ray crystal structure data of the Torpedo californica acetylcholinesterase (Sussman, J.L., Harel, M., Frolow, F., Oefner, C., Goldman, A., Toker, L. and Silman, I. (1991) Science 253, 872-879). Attempts to convert the AChE triad into a Cys-His-Glu or Ser-His-Asp configuration by site-directed mutagenesis did not yield effective AChE activity. Another type of substitution, that of Asp-74 by Gly or Asn, generated an active enzyme with increased resistance to succinylcholine and dibucaine; thus mimicking in an AChE molecule the phenotype of the atypical butyrylcholinesterase natural variant (D70G mutation). Mutations of other carboxylic residues Glu-84, Asp-95, Asp-333, and Asp-349, all conserved among cholinesterases, did not result in detectable alteration in the recombinant AChE, although polypeptide productivity of the D95N mutant was considerably lower. In contrast, complete absence of secreted human AChE polypeptide was observed when Asp-175 or Asp-404 were substituted by Asn. These two aspartates are conserved in the entire cholinesterase/thyroglobulin family and appear to play a role in generating and/or maintaining the folded state of the polypeptide. The x-ray structure of the Torpedo acetylcholinesterase supports this assumption by revealing the participation of these residues in salt bridges between neighboring secondary structure elements.
ESTHER : Shafferman_1992_J.Biol.Chem_267_17640
PubMedSearch : Shafferman_1992_J.Biol.Chem_267_17640
PubMedID: 1517212

Title : Substrate inhibition of acetylcholinesterase: residues affecting signal transduction from the surface to the catalytic center - Shafferman_1992_EMBO.J_11_3561
Author(s) : Shafferman A , Velan B , Ordentlich A , Kronman C , Grosfeld H , Leitner M , Flashner Y , Cohen S , Barak D , Ariel N
Ref : EMBO Journal , 11 :3561 , 1992
Abstract : Amino acids located within and around the 'active site gorge' of human acetylcholinesterase (AChE) were substituted. Replacement of W86 yielded inactive enzyme molecules, consistent with its proposed involvement in binding of the choline moiety in the active center. A decrease in affinity to propidium and a concomitant loss of substrate inhibition was observed in D74G, D74N, D74K and W286A mutants, supporting the idea that the site for substrate inhibition and the peripheral anionic site overlap. Mutations of amino acids neighboring the active center (E202, Y337 and F338) resulted in a decrease in the catalytic and the apparent bimolecular rate constants. A decrease in affinity to edrophonium was observed in D74, E202, Y337 and to a lesser extent in F338 and Y341 mutants. E202, Y337 and Y341 mutants were not inhibited efficiently by high substrate concentrations. We propose that binding of acetylcholine, on the surface of AChE, may trigger sequence of conformational changes extending from the peripheral anionic site through W286 to D74, at the entrance of the 'gorge', and down to the catalytic center (through Y341 to F338 and Y337). These changes, especially in Y337, could block the entrance/exit of the catalytic center and reduce the catalytic efficiency of AChE.
ESTHER : Shafferman_1992_EMBO.J_11_3561
PubMedSearch : Shafferman_1992_EMBO.J_11_3561
PubMedID: 1396557

Title : Molecular Models for Human AChE and its Phosphonylation Products -
Author(s) : Barak D , Ariel N , Velan B
Ref : In Multidisciplinary approaches to cholinesterase functions - Proceedings of Fourth International Meeting on Cholinesterases , (Shafferman, A. and Velan, B., Eds) Plenum Press, New York :195 , 1992
PubMedID:

Title : Serologically defined linear epitopes in the E2 envelope glycoprotein of Semliki Forest virus - Ariel_1990_Arch.Virol_113_99
Author(s) : Ariel N , Lehrer S , Elhanaty E , Sabo T , Brodt P , Lachmi B , Katz D , Levin R , Grosfeld H , Velan B , et al.
Ref : Arch Virol , 113 :99 , 1990
Abstract : A set of 41 overlapping peptides, representing the complete sequence of SFV-E2 protein were synthesized and analyzed in the ELISA test against murine anti-SFV sera. No single peptide was recognized by all antisera. Eight peptides were found to be highly reactive with hyperimmune anti-SFV sera. Six out of the eight peptide sequences coincide with the most hydrophilic regions of SFV-E2. Out of these, four peptides (amino acid positions 16-35, 61-80, 166-185, 286-305) that contain the least number of alphavirus conserved residues were selected. This panel constitutes the minimal number of peptides necessary and sufficient for specific recognition of hyperimmune mouse anti-SFV sera.
ESTHER : Ariel_1990_Arch.Virol_113_99
PubMedSearch : Ariel_1990_Arch.Virol_113_99
PubMedID: 1696808