Imidazole Binding Research Articles

Azide, cyanide, fluoride, imidazole and pyridine binding to ferric and ferrous native horse heart cytochrome c and to its carboxymethylated derivative: A comparative study

Azide, cyanide, fluoride, imidazole, and pyridine binding to ferric and ferrous native horse heart cytochrome c and to its carboxymethylated derivative has been investigated, from the thermodynamic viewpoint, at pH 7.5 and 25.0°C. Ligand affinity for ferric and ferrous carboxymethylated cytochrome c is higher by about 30- and 400-fold, respectively, than that observed for the native protein. The results here reported: (i) allow the estimation, for the first time, of the ligand-independent free energy associated with the heme-iron sixth coordination bond in ferric and ferrous native cytochrome c, which turns out to be +8.4 kJ mol −1 and +14.6 kJ mol −1, at 25.0°C, respectively, and (ii) suggest an interplay between redox, structural, ligand binding, and recognition properties of cytochrome c.

Kinetic mechanism of folding and unfolding of Rhodobacter capsulatus cytochrome c2.

In spite of marginal sequence homology, cytochrome c2 from photosynthetic bacteria and the mitochondrial cytochromes c exhibit some striking structural similarities, including the tertiary arrangement of the three main helices. To compare the folding mechanisms for these two distantly related groups of proteins, equilibrium and kinetic measurements of the folding/unfolding reaction of cytochrome c2 from Rhodobacter capsulatus were performed as a function of guanidine hydrochloride (GuHCl) concentration in the absence and presence of a stabilizing salt, sodium sulfate. Quenching of the fluorescence of Trp67 by the heme was used as a conformational probe. Kinetic complexities due to non-native histidine ligation are avoided, since cytochrome c2 contains only one histidine, His17, which forms the axial heme ligand under native and denaturing conditions. Quantitative kinetic modeling showed that both equilibrium and kinetic results are consistent with a minimal four-state mechanism with two sequential intermediates. The observation of a large decrease in fluorescence during the 2-ms dead-time of the stopped-flow measurement (burst phase) at low GuHCl concentration, followed by a sigmoidal recovery of the initial amplitude toward the unfolding transition region, is attributed to a well-populated compact folding intermediate in rapid exchange with unfolded molecules. A nearly denaturant-independent process at low GuHCl concentrations reflects the rate-limiting conversion of a compact intermediate to the native state. At high GuHCl concentrations, a process with little denaturant dependence is attributed to the rate-limiting Met96-iron deligation process during unfolding, which is supported by the kinetics of imidazole binding. The strong GuHCl-dependence of folding and unfolding rates near the midpoint of the equilibrium transition is attributed to destabilization of each intermediate and their transition states in folding and unfolding. Addition of sodium sulfate shifts the rate profile to higher denaturant concentration, which can be understood in terms of the relative stabilizing effect of the salt on partially and fully folded states.

Molecular modelling of inhibitors of 17α-Hydroxylase — a novel approach

A novel molecular modelling study, involving inhibitors bound to a ‘substrate-heme complex’, is described for the binding of non-steroidal imidazole based inhibitors of the 17α-Hydroxylase (17α-OHase) component of the enzyme complex 17α-Hydroxylase/17,20-Lyase to gain insight into the active site of the overall enzyme.

Nonadrenergic binding of [3H]atipamezole in rat lung. A novel imidazole binding site?

Nonadrenergic binding of [3H]atipamezole in rat lung. A novel imidazole binding site?

Construction of a bisaquo heme enzyme and binding by exogenous ligands.

The crystal structure of the His-175-->Gly (H175G) mutant of cytochrome-c peroxidase (EC 1.11.1.5), missing its only heme ligand, reveals that the histidine is replaced by solvent to give a bisaquo heme protein. This protein retains some residual activity, which can be stimulated or inhibited by addition of exogenous ligands. Structural analysis confirms the binding of imidazole to the heme at the position of the wild-type histidine ligand. This imidazole complex reacts readily with hydrogen peroxide to produce a radical species with novel properties. However, reactivation in this complex is incomplete (approximately 5%), which, in view of the very similar structures of the wild-type and the H175G/imidazole forms, implies a critical role for tethering of the axial ligand in catalysis. This study demonstrates the feasibility of constructing heme enzymes with no covalent link to the protein and with unnatural ligand replacements. Such enzymes may prove useful in studies of electron transfer mechanisms and in the engineering of novel heme-based catalysts.

Application Of 2D Exchange Spectroscopy to the Studies of Imidazole Binding to CytochromeC

Abstract Two-dimensional 1H exchange spectroscopy has been applied to the studies of imidazole binding to cytochrome c. The magnetization exchange rate constants, the substitution reaction rate constants and the equilibrium constants have been evaluated. Some resonances of imidazole substituted forms (Im-cyt c) have been assigned by 2D exchange NMR. The observed hyperfine shifts of heme methyl groups of Im-cyt c and cyt c have been corrected for contact and dipolar contributions. The detailed analysis of these results indicates that the axial ligands and their orientations, and the substituents of heme are important determinants of the electronic structure of the heme in cyt c and its derivatives.

Selective Inhibition of Mammalian Lanosterol 14.alpha.-Demethylase by RS-21607 in vitro and in vivo

The discovery of selective lanosterol 14 alpha-demethylase inhibitors may lead to novel hypolipidemic drugs. RS-21607, (2S,4S)-cis-2[1H-imidazol-1-yl)methyl]-2-[2-(4-chlorophenyl)ethyl]-4- [[(4-aminophenyl)thio]methyl]-1,3-dioxolane, was characterized as a tight-binding, competitive inhibitor of lanosterol 14 alpha-demethylase purified from rat liver. The apparent Ki was determined to be 840 pM and found to be similar in hepatic microsomes from human, rat, and hamster. RS-21607, which contains two chiral centers, was a more effective lanosterol 14 alpha-demethylase inhibitor than its three stereoisomers. In vitro, RS-21607 had a greater affinity for lanosterol 14 alpha-demethylase than the other cytochromes P450 evaluated: CYP7, CYP27, CYP11A1, CYP19, CYP17, CYP11B1, CYP21, CYP3A4, CYP4A, CYP2D6, CYP1A2, CYP2C9, and 27-hydroxycholesterol 7 alpha-hydroxylase. The other stereoisomers were not as selective as RS-21607. Doses of 3-30 mg/kg RS-21607 given orally to hamsters caused a dose-dependent decrease in cholesterol biosynthesis with a corresponding accumulation of 24,25-dihydrolanosterol. RS-21607 inhibited the enzyme and cholesterol biosynthesis in hamster liver by 50% at 18 h following a 30 mg/kg oral dose. This was interpreted to indicate that RS-21607 is able to distribute to the site of action in hamsters and inhibit the target enzyme. In the same dose range, the plasma concentrations of testosterone, corticosterone, and progesterone, the endpoints for the cytochromes P450 involved in steroid biosynthesis, were relatively unaffected. These data show RS-21607 to be an effective and selective inhibitor of lanosterol 14 alpha-demethylase, both in vivo and in vitro. RS-21607 interacted with the purified enzyme to produce a type II binding spectrum, consistent with an interaction between the imidazole moiety and the heme. The electrostatic contribution of the imidazole binding was investigated using the desimidazole analog of RS-21607. The apparent Ki for the desimidazole compound (65 microM) was similar to the apparent Km for the substrate DHL (79 microM). Together, these data confirm that the ligand attached to the imidazole in RS-21607 is a good non-sterol substitute for DHL, i.e., binding to the enzyme with similar affinity, and that the coordination of the imidazole to the heme provides a major electrostatic contribution for the inhibition of lanosterol 14 alpha-demethylase by RS-21607. RS-21607 was also observed to increase the accumulation of 3 beta-hydroxy-24,25-dihydrolanost-8-en-32-al, the second intermediate in the multistep oxidation, but not the first intermediate. 24,25-dihydrolanost-8-ene-3 beta,32-diol.(ABSTRACT TRUNCATED AT 400 WORDS)

Small Molecule Binding to an Artificially Created Cavity at the Active Site of Cytochrome c Peroxidase

In the oxidized "ES" state of cytochrome c peroxidase, Trp-191 is reversibly oxidized to a stable cation free radical by the hypervalent heme. To explore the potential for engineering a binding site for heterocyclic compounds at this site, the mutant W191G was constructed. Two independent crystal structures of W191G at 2.1- and 2.3-A resolution show that W191G contains a well-defined, approximately 180-A3 cavity at the Trp-191 site. The cavity is occupied by five ordered water molecules which participate in an extensive hydrogen-bonding network with each other, with polar main-chain atoms, and with the carboxylate of Asp-235. After a number of heterocyclic compounds were screened, evidence was obtained that substituted imidazoles bind to the cavity of W191G. Titration of W191G with imidazole resulted in a perturbation of the Soret absorption band that was not observed for W191H, W191F, or the native enzyme. The dissociation constants for binding of benzimidazole, imidazole, 2-ethylimidazole, 1-methylimidazole, 2-methylimidazole, and 1,2-dimethylimidazole to W191G were respectively 2.58, 0.70, 0.36, 0.057, 0.047, and 0.027 mM at pH 6.0. The highest binding affinity was exhibited by 1,2-dimethylimidazole, indicating that steric interactions and the efficiency of filling the cavity are important determinants for specificity. The Kd for imidazole binding increased from 0.7 mM at pH 6 to 3.0 mM at pH 8 and could be fit to a single proton ionization curve with a pKa of 7.4, demonstrating the preferential binding by the imidazolium ion (pKa = 7.3). The binding of a number of substituted imidazoles to the cavity of W191G was verified by X-ray crystallographic analysis. The most clearly defined density was observed for W191G crystals soaked in 1 mM 1,2-dimethylimidazole and was consistent with an oriented occupation in which the unsubstituted nitrogen forms a hydrogen bond or ion pair interaction with Asp-235. Thus, enhanced binding of positively charged molecules may be the result of interactions with this carboxylate. An analogous interaction may stabilize the developing positive charge on the Trp-191 radical of the wild-type enzyme. While the oxidation of imidazoles by the ferryl intermediate of W191G was neither expected nor observed, this study has defined the structural determinants for small molecule binding to an artificially created cavity near a heme center which is capable of generating oxidized species at a potential of over 1 V, and these results will guide future attempts for novel substrate oxidation by CCP.

Thermodynamics of binding of the CO2-competitive inhibitor imidazole and related compounds to human carbonic anhydrase I: an isothermal titration calorimetry approach to studying weak binding by displacement with strong inhibitors.

The visible spectrum of Co(II)-substituted human carbonic anhydrase I (HCA I) complexed with the unique CO2-competitive inhibitor imidazole undergoes a marked alkaline intensification, with a midpoint near pH 8 [Bauer, R., Limkilde, P., & Johansen, J. T. (1977) Carlsberg Res. Commun. 42, 325-339]. This change was first attributed to the ionization of a nondisplaced water ligand of the active-site metal in a five-coordinate complex. Later proposals favored assigning it to the deprotonation of the bound imidazole itself to give a tetrahedrally coordinated imidazolate anion at high pH. We have determined by isothermal titration calorimetry the pH dependence of the enthalpy of binding of imidazole and its analogues to HCA I and Co(II)HCA I. We devised an indirect strategy whereby the enthalpy of binding of the strong sulfonamide inhibitor methazolamide was determined in the absence and presence of a constant high concentration of the competing imidazole or its analogues. The standard enthalpy of binding of deprotonated methazolamide to the "acid" form of HCA I and Co(II)HCA I was found to be pH independent over the pH range of 6.5-9.5, as expected. It was also identical for both the zinc (-13.5 +/- 1.1 kcal M-1) and the cobalt (-13.7 +/- 0.4 kcal M-1) forms. The standard enthalpy of binding of neutral imidazole (average value -6.1 +/- 0.8 kcal M-1) surprisingly did not show any marked pH dependence, varying by about 1.1 and 2.6 kcal M-1 for the zinc and cobalt enzymes, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Haem–peptide complexes. Synthesis and stereoselective oxidations by deuterohaemin-L-phenylalanyl-poly-L-alanine complexes

Deuterohaemin-L-histidine methyl ester—peptide complexes have been obtained by covalently linking L-histidine methyl ester and the peptides Ala-Ala-Phe-Ala-Ala-Ala-Ala-Ala-Ala-Ala (compound 3) or Ala-Ala-Ala-Phe-Ala-Ala-Ala-Ala-Ala-Ala (compound 4) on the propionic acid side chains. The spectroscopic properties of 3 and 4 and the imidazole binding equilibria indicate that folding of the peptide chain on the opposite part of the porphyrin plane with respect to that occupied by the bound histidine side-arm reduces in the order 3 > 4 the accessibility of exogenous ligands to the sixth co-ordination position of the iron atom, probably through some stacking interactions between the phenylalanine residue of the peptide and the porphyrin ring in the case of 3. This arrangement has marked consequences on the stereoselectivity observed in a model peroxidase reaction using L- or D-tyrosine methyl ester as substrates and tert-butyl hydroperoxide as oxidant in dichloromethane–trifluoroethanol (9:1), that have been interpreted in terms of the interaction between the chiral substrates and the peptide chains of the deuterohaemin complexes.

Imidazole binding sites in rabbit kidney and forebrain membranes

1. The binding of [3H]-clonidine, [3H]-idazoxan and [3H]-yohimbine to rabbit forebrain and kidney membranes was compared. 2. Yohimbine bound exclusively to adrenergic sites, idazoxan to non-adrenergic sites and clonidine to both non-adrenergic and adrenergic sites. 3. Differences were observed between the ligands not only in binding at adrenergic and non-adrenergic sites but also between the non-adrenergic binding of [3H]-clonidine and [3H]-idazoxan. 4. However, no tissue specific differences were found.

Rilmenidine lowers arterial pressure via imidazole receptors in brainstem C1 area

We sought to determine the site of action and receptor type responsible for the antihypertensive actions of rilmenidine. an oxazoline analogue of clonidine. In anesthetized paralyzed rats decerebration did not alter the dose dependent reductions in arterial pressure and heart rate elicited by i.v. drug. Rilmenidine microinjected bilaterally into the C1 area of the rostral ventrolateral medulla (RVL), but not nucleus tractus solitarii (NTS) nor caudal ventrolateral medulla (CVL), elicited dose-dependent falls in arterial pressure and heart rate at doses an order of magnitude less than required systemically. Prior microinjection into the Cl area of the selective α 2-adrenoceptor antagonist SKF-86466, even at high doses, failed to modify the hypotension to i.v. rilmenidine. However, microinjection of 3- to 10-fold lower doses of idazoxan, a ligand for imidazole as well as α 2-adrenoceptors, blocked the effects. Rilmenidine also competed with the clonidine analogue [ 3H]p-aminoclonidine ([ 3H]pac) at specific binding sites in membranes of bovine ventrolateral medulla and frontal cortex. In RVL rilmenidine competed with binding to imidazole and α 2-adrenergic binding sites with a 30-fold selectivity for the imidazole binding sites. In frontal cortex binding was of lower affinity and restricted to α 2-adrenergic sites. We conclude that rilmenidine, like clonidine, acts to lower arterial pressure by an action on imidazole receptors in the Cl area of RVL. The higher selectivity of rilmenidine for imidazole to α 2-adrenoceptors as compared to clonidine may explain the lower sedative effects of rilmenidine. Rilmenidine; Oxazoline; α 2-Adrenoceptors; Imidazole receptors; Medulla (rostral ventrolateral); Arterial pressure

Down-regulation of α 2 adrenoceptors in ventrolateral medulla of spontaneously hypertensive rats

The binding of [ 3H]idaxazon to imidazole sites and [ 3H]rauwolscine to α adrenoceptors of neuronal membranes prepared from cerebral cortex and ventrolateral medulla of 10 week old spontaneously hypertensive (SHR) and normotensive Wistar-Kyoto (WKY) rats was determined. [ 3H]idaxazon bound to the membranes of cerebral cortex and ventrolateral medulla at a single high affinity site. The binding of [ 3H]idaxazon in ventrolateral medulla and cerebral cortex was found to be similar in SHR and WKY rats. [ 3H]Rauwolscine bound to the membranes of cerebral cortex and ventrolateral medulla at a single high affinity site. The binding of [ 3H]rauwolscine in the cerebral cortex was found to be similar in SHR and WKY rats. However, in the ventrolateral medulla [ 3H]rauwolscine binding was found to be significantly lower in SHR as compared to WKY rats. The decreased binding was due a decrease (32%) in the B max value in SHR rats as compared to WKY rats. The K d values were similar in SHR and WKY rats. It is concluded that imidazole binding sites are not affected while, α 2 adrenergic binding sites are decreased in the ventrolateral medulla of SHR rats and may be contributing to the regulation of blood pressure.

13C and 1H NMR studies of imidazole binding to native and Co(II)-substituted human carbonic anhydrase I

The inhibition constant of imidazole toward human carbonic anhydrase I has been directly determined at pH 6.8 and 8.8 through 13C NMR. The data are in agreement with those obtained by indirect methods and confirm that the binding affinity of imidazole is substantially constant in the pH range investigated. Analysis of the isotropically-shifted signals in the 1H NMR spectra of the Co(II)-substituted enzyme interacting with imidazole at high and low pH in the presence of sulfate as counterion indicates the likely formation of a five-coordinate HCAI-imidazole-sulfate adduct at low pH. The binding mode of imidazole in the active site of the enzyme is discussed.

Kinetic studies of the molecular aggregation of Limulus oxyhemocyanin and apohemocyanin

The kinetic dissociation of Limulus oxyhemocyanin and apohemocyanin in alkaline pH have been studied using a high-performance liquid chromatographic method capable of measuring both the subunit and the aggregated proteins. The results show that the oxy- and apohemocyanins exhibit two distinct dissociation kinetics marked by their differences in aggregation patterns. For oxyhemocyanin, the kinetics occur at pH 7.8 and above and show a three-step dissociation of the 24-mer to the subunits. Each reaction step is characterized by the ionization of one set of protons with higher p K a associated with the dissociation of smaller aggregates. Binding of imidazole at or near the copper active site inhibits the dissociation kinetics. Cyanide ion, which removes the active copper from the protein, causes complete dissociation of the protein into subunits. For apohemocyanin, a two-step dissociation of the protein into subunits at pH 7.6 and above was observed. The kinetic results show a subunit ratio of 3:2 for the two aggregates, suggesting that the subunits of the copper-free protein may aggregate into dimers and trimers. In the pH range 6.5–7.5, apohemocyanin does not undergo kinetic dissociation. Since the protein has the highest state of aggregation at pH 6.5, association of the trimers into a larger aggregate such as a hexamer at this pH is also postulated.

Characterization and Visualization of Clonidine-sensitive Imidazole Sites in Rat Kidney Which Recognize Clonidine-displacing Substance

In the ventrolateral medulla oblongata, clonidine binds not only to alpha 2-adrenergic receptors but also to a novel class of nonadrenergic sites that are specific for imidazoles. Since clonidine has direct actions on the kidney, we sought to determine whether imidazole binding sites could be detected in renal cell membranes. Adrenergic agents having an imidazole ring, like clonidine, completely displaced the specific binding of the high-affinity clonidine analog 3H-p-aminoclonidine (3H-PAC) to kidney membranes. Nonimidazole adrenergic agents inhibited 3H-PAC binding by only 75%, indicating that the remaining 25% of the sites were nonadrenergic. Cimetidine, an imidazole compound lacking adrenergic potency, showed a high affinity for approximately 25% of the sites. 3H-PAC binding to imidazole sites was high-affinity (KD = 11 +/- 3 nmol/L) and saturable (Bmax = 41 +/- 10 fmol/mg protein). Like clonidine, an endogenous clonidine-displacing substance (CDS) completely inhibited 3H-PAC binding to renal cortex membranes. Quantitative receptor autoradiography revealed that imidazole receptors exhibited a specific regional distribution within the kidney that was unique, and distinct from that of alpha 2-adrenergic receptors. We conclude that clonidine binds to specific imidazole sites in the renal cortex of the rat and that CDS may be the endogenous ligand at these sites. Thus, in addition to their role in central nervous system control of arterial pressure, imidazole receptors may be involved in the regulation of renal function.

Equilibrium and kinetic study of imidazole binding to phthalocyaninatoiron(II) in dimethyl sulphoxide

Equilibrium and kinetic study of imidazole binding to phthalocyaninatoiron(II) in dimethyl sulphoxide

Inhibition of β-glucosidase by imidazoles

Over 25 nitrogen-containing heterocycles were tested as inhibitors of sweet almond beta-glucosidase (EC 3.2.1.21). Among the most potent of these are some imidazole derivatives. The pH dependence indicates that the unprotonated inhibitor binds most tightly to the catalytically active species of the enzyme. This is analogous to the situation with 1-deoxynojirimycin where the permanently cationic species, N,N-dimethyl-1-deoxynojirimycin, binds at least two orders of magnitude less tightly to the enzyme than does the unprotonated 1-deoxynojirimycin. The binding of imidazole derivatives show a general tendency of increasing affinity with increasing basicity (beta approximately 0.4). One derivative which shows a significant positive deviation from this correlation (- log Ki vs. pKa) is 4-phenylimidazole. 4-Phenylimidazole is one of the most potent reversible inhibitors of beta-glucosidase with a pH-independent Ki = 0.8 microM. It is also fairly specific for beta-glucosidase, binding at least three orders of magnitude less tightly to any of the other exoglycosidases tested. This inhibitor combines, in a mono-molecular species, the binding affinities of benzene, which binds at the hydrophobic aglycone binding site, and imidazole, which binds at the sugar binding site of beta-glucosidase. The binding energy of 4-phenylimidazole can be attributed to the sum of the intrinsic binding energies of the phenyl and imidazole moieties. Thus, there is no significant entropic advantage of combining the component parts of phenylimidazole in a single species. This indicates that there is no significant uncompensated entropy loss upon binding of either benzene or imidazole to the enzyme. Nevertheless, the additivity of binding energy, even in the absence of an entropic advantage, results in the most powerful known inhibitor of the enzyme.

Protein-protein interactions in microsomal cytochrome P-450 isozyme LM2 and their effect on substrate binding.

The effects of protein-protein interactions and substrate binding on the structure of the active site of rabbit liver microsomal cytochrome P-450 LM2 have been analyzed by resonance Raman spectroscopy of the monomeric and oligomeric protein in solution. Also H2O2-dependent catalytic activities of the two states have been compared. The two vinyl substituents of the heme exhibit different orientations, as indicated by the frequencies and intensities of their stretching vibrations. One group lies in the plane of the heme and remains unchanged in the two states of cytochrome P-450 LM2, the other is tilted out of the plane. The tilting angle in oligomers was smaller than in monomers. These vinyl stretching modes together with some porphyrin modes, were found to be sensitive indicators of the quaternary structure and of substrate binding. In both the oligomer and the monomer, substrate binding causes changes of the relative intensities of some porphyrin modes and the vinyl stretching vibrations which may reflect modifications of the electronic transitions due to hydrophobic interactions between the bound substrate and the heme. In contrast to the monomeric cytochrome P-450 LM2, benzphetamine binding to the oligomers of this isozyme additionally produces a shift of the spin-state equilibrium. This indicates that in the oligomer the substrate-binding pocket is converted by protein-protein interaction to a structure that forces substrates to interfere with the sixth ligands, inducing an increase of the five-coordinated high-spin configuration. In the monomer the substrate-binding pocket can accommodate benzphetamine without affecting the spin state. Binding of imidazole to the monomeric and oligomeric cytochrome P-450 LM2 produces essentially the same resonance Raman spectra. Apparently the replacement of the native sixth ligand by imidazole disturbs the structure of the active site in such a way that it becomes insensitive to protein-protein interactions. H2O2-dependent N-demethylation of benzphetamine and aniline p-hydroxylation by cytochrome P-450 LM2 did not depend on its state of aggregation.

ChemInform Abstract: Selective Binding of Imidazoles and Related Organic Molecules in an Organic Solvent

ChemInformVolume 20, Issue 28 Reviews ChemInform Abstract: Selective Binding of Imidazoles and Related Organic Molecules in an Organic Solvent J. JUN. REBEK, J. JUN. REBEK Univ. Pittsburgh, Pittsburgh, PA, USASearch for more papers by this author J. JUN. REBEK, J. JUN. REBEK Univ. Pittsburgh, Pittsburgh, PA, USASearch for more papers by this author First published: July 11, 1989 https://doi.org/10.1002/chin.198928349Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinked InRedditWechat No abstract is available for this article. Volume20, Issue28July 11, 1989 RelatedInformation