β-Galactosidase (EC 3.2.1.23) is known to be inhibited by some thiol reagents. 1-Benzoyl-1-cyano-2-(4,5-dimethoxy-2-nitrophenyl)-ethene (1) was shown to be an irreversible inhibitor, while 1, 1-dicyano-2-(4,5-dimethoxy-2-nitrophenyl)-ethene (2) was demonstrated as a positive irreversible modulator causing a rise of up to 186% in β-galactosidase activity. Compound 2 is, however, an irreversible inhibitor of the cysteine proteinase papain (preceding paper). Kinetic values of β-galactosidase at pH 8.3 with o-nitrophenyl β-D-galactopyranoside (ONPG) as the substrate and for compounds 1 and 2 were determined and in view of model experiments, it was assumed that both compounds possibly reacted with the thiol side chain of Cys in the active site inducing allosteric changes in the enzyme. Since the enzyme, modified by compound 1 or 2, was a 2-nitrobenzyl derivative, near-UV irradiation resulted in a recovery of up to 91% and a reduction of the enzyme's activity to 90%, respectively.

1-(4,5-Dimethoxy-2-nitrophenyl)-2-nitroethene (1) was shown to be an irreversible inhibitor of papain (EC 3.4.22.2), causing a complete inhibition (120 min preincubation, pH 8.0), assuming that it attached to Cys-25 at the active site of the enzyme (while a short preincubation time caused activation). Only partial inhibition of papain was achieved, however, with 1,1-dicyano-2-(4,5-dimethoxy-2-nitrophenyl)-ethene (2), a compound synthesized in this work, which is also an irreversible inhibitor of papain. Since both compounds 1 and 2, and in each case of the inhibited enzyme, were 2-nitrobenzyl derivatives, they and the modified enzyme were expected to be photosensitive. Indeed, irradiation of the inhibited enzyme in the presence of mercaptoethanol resulted in a full recovery of the enzyme activity following inactivation with compound 1 (similar to our previous finding with β-galactosidase) and up to 67% recovery following inhibition with compound 2.

We demonstrate the utility of the new principle of Universal Standard Reagents (USR) in glycosylation of protected sugars and selective hydrolysis of ester glycosides by means of 3-(2,4-dinitroanilinopropanol) (DNAP) and 1-(2,4-dinitropheny1)-4-hydroxypiperidine (DNPP).Quantitative determination of each individual sugar derivative was carried out using extinction coefficients at A , ,(-350 nm) on the micromole scale, eliminating the need for specific standards.The selective hydrolysis of P-DNAP-tetra-0-acetyl-and 0aroylglycosides was examined by different 0-and N-bases using quantitative assessment by TLC and HPLC.DCIiNH3 MS of anomers and NMR-spectra of partially acylated glucosides are discussed.

4-(Phenylethynyl)-6-phenyl-1,4-dihydropyridine derivatives are selective antagonists at human A3 adenosine receptors, with Ki values in a radioligand binding assay vs [125I]AB-MECA (N6-(4-amino-3-iodobenzyl)-5'-(N-methylcarbamoyl)adenosine) in the submicromolar range. In this study, structure-activity relationships at various positions of the dihydropyridine ring (the 3- and 5-acyl substituents, the 4-aryl substituent, and 1-methyl group) were probed synthetically. Using the combined protection of the 1-ethoxymethyl and the 5-[2-(trimethylsilyl)ethyl] ester groups, a free carboxylic acid was formed at the 5-position allowing various substitutions. Selectivity of the new analogues for cloned human A3 adenosine receptors was determined vs radioligand binding at rat brain A1 and A2A receptors. Structure-activity analysis at adenosine receptors indicated that pyridyl, furyl, benzofuryl, and thienyl groups at the 4-position resulted in, at most, only moderate selectivity for A3 adenosine receptors. Ring substitution (e.g., 4-nitro) of the 4-phenylethylnyl group did not provide enhanced selectivity, as it did for the 4-styryl-substituted dihydropyridines. At the 3-position of the dihydropyridine ring, esters were much more selective for A3 receptors than closely related thioester, amide, and ketone derivatives. A cyclic 3-keto derivative was 5-fold more potent at A3 receptors than a related open-ring analogue. At the 5-position, a homologous series of phenylalkyl esters and a series of substituted benzyl esters were prepared and tested. (Trifluoromethyl)-, nitro-, and other benzyl esters substituted with electron-withdrawing groups were specific for A3 receptors with nanomolar Ki values and selectivity as high as 37000-fold. A functionalized congener bearing an [(aminoethyl)amino]carbonyl group was also prepared as an intermediate in the synthesis of biologically active conjugates.

1-Nitro-2-phenylethene (β-nitrostyrene, 1), which is a thiol-protecting reagent (Jung, G., Fouad, H. and Heusel, G. (1975) Angew. Chem. Int. Ed. Engl. 14, 817–818), was demonstrated in this work to be an irreversible inhibitor of β-galactosidase (EC 3.2.1.23), an enzyme known to be inhibited by some thiol reagents or though modifying a methionine residue at the active site. No reversal of the inhibition was observed upon subsequent incubation with mercaptoethanol or irradiation (350 nm). 1-(4,5-dimethoxy-2-nitrophenyl)-2-Nitroethene (2) was also shown to be an irreversible inhibitor (94% inhibition, pH 8.3) of the enzyme. Kcat values of β-galactosidase at pH 8.3 with o-nitrophenyl β-d-galactopyranoside (ONPG) as the substrate and at the highest inhibitor concentrations employed for compound 1 (4.06 · 10−4 M) ranged from 1.67 · 104 s−1 after 30 min of preincubation to < 0.07 · 104 s−1 after 180 min preincubation. For compound 2 (9.5 · 10−5 M) Kcat values ranged from 2.70 · 104 s−1 following 30 min preincubation to 1.15 · 104 s−1 after 180 min of preincubation; the changes in Kmapp, however, were small. The activity was not recovered following incubation with mercaptoethanol. Since compound 2 and the inhibited enzyme are 2-nitrobenzyl derivatives, they are expected to be photosensitive and indeed, irradiation of the inhibited enzyme in the presence of mercaptoethanol resulted in recovery (89%, pH 8.3) of the enzyme activity.

Makromolekulare Chemie. Macromolecular SymposiaVolume 70-71, Issue 1 p. 455-457 Article Novel uses of polymeric reagents in chemical transformation A. Patchornik, A. Patchornik The Weizmann Institute of Science, Rehovot, and Bar-Ilan University, Ramat Gan, IsraelSearch for more papers by this author A. Patchornik, A. Patchornik The Weizmann Institute of Science, Rehovot, and Bar-Ilan University, Ramat Gan, IsraelSearch for more papers by this author First published: May 1993 https://doi.org/10.1002/masy.19930700144AboutPDF 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 Volume70-71, Issue1May 1993Pages 455-457 RelatedInformation

A. Patchornik, Y. Ben-David and D. Milstein, J. Chem. Soc., Chem. Commun., 1990, 1090 DOI: 10.1039/C39900001090

The use of transfer polymeric reagents (PRs) as excellent acylating agents for high yield and high purity peptide synthesis is described. Three methodologies are compared: the classical solution method, the Merrifield approach and an automated (the "mediator"-shadchan) method with continuous monitoring. The utilization of PRs as general acyl transfer reagents is also elaborated. The described approaches are not limited to peptide synthesis, but may be applicable to a wide range of organic reaction types.

We propose a novel concept of photo reversible affinity labeling (ReAL) in which the molecule is labeled by the formation of a photosensitive linkage and is later cleaved under mild physiological conditions by the action of light.

Chemischer InformationsdienstVolume 17, Issue 48 Reviews ChemInform Abstract: Polymeric Transfer Reagents for Organic Synthesis with Self-Control toward Automation in Organic Synthesis A. PATCHORNIK, A. PATCHORNIKSearch for more papers by this authorE. NOV, E. NOVSearch for more papers by this authorK. A. JACOBSON, K. A. JACOBSONSearch for more papers by this authorY. SHAI, Y. SHAISearch for more papers by this author A. PATCHORNIK, A. PATCHORNIKSearch for more papers by this authorE. NOV, E. NOVSearch for more papers by this authorK. A. JACOBSON, K. A. JACOBSONSearch for more papers by this authorY. SHAI, Y. SHAISearch for more papers by this author First published: December 2, 1986 https://doi.org/10.1002/chin.198648389Read 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 onEmailFacebookTwitterLinkedInRedditWechat No abstract is available for this article. References A. PATCHORNIK, E. NOV, K. A. JACOBSON, Y. SHAI, Polymeric Transfer Reagents for Organic Synthesis with Self-Control toward Automation in Organic Synthesis, ACS Symp. Ser., 1986, 308, 231. Volume17, Issue48December 2, 1986 ReferencesRelatedInformation

AbstractEine neue Methodologie für eine potentiell automatische selbststeuernde Synthese von Peptiden wird beschrieben.

A synthetic strategy to obtain chelating polymers via a one-step functionalization of selected macromolecular networks, such as macroporous styrene-divinylbenzene copolymers, is presented. The conversion of chloromethylpolystyrene to aminomethylpolystyrene in a Delepine reaction by way of hexamethylenetetramine addition to the chloride, followed by acid hydrolysis, provides a nucleophilic polymer of high activity. It can be alkylated with a variety of disubstituted benzylhalides (2-acyl-4-chloromethylphenols, 3-picolylchloride, 5-chloromethyl-8-hydroxyquinoline) to yield chelating polymers with strong affinity for transition metal ions. The metal complexing properties of the polymers agree well with estimated properties from known K1 values of the parent ligands in homogenous media. The chelating polymers, 5A, 6A and 7A show very good reversibility in metal binding and release, and good kinetic behaviour due to the hydrophilic nature of the -CH2NHCH2-spacer group between the backbone and the chelating group.

A series of highly colored nitrophenolates and nitrothiophenolates has been tested as spray reagents for the detection of electrophilic species of the types commonly used in peptide and protein chemistry. Sensitive TLC detection of agents for alkylation, acylation, sulfonylation and phosphorylation was demonstrated. In addition, the thiophenolate sprays were sensitive for oxidizing agents in nanomolar quantities. Selective TLC detection of acylating and phosphorylating agents was accomplished by subsequent alkali treatment resulting in the restoration of color.

In this report we further show the utility and efficiency of polymer-bound 1-hydroxybenzotriazole (PHBT) as an almost ideal support for the polymeric reagent method of peptide synthesis. This was demonstrated by the synthesis of thymosin alpha 1 (15-28), in which two suitably blocked segments, Boc-Asp (OtBu)-Leu-Lys (2Cz)-Glu (OBzl)-Lys (2Cz)-Lys (2Cz)-OH (3) and Boc-Glu (OBzl)-Val-Val-Glu (OBzl)-Glu (OBzl)-Ala-Glu (OBzl)-Asn-OBzl (2), were prepared entirely by utilizing PHBT activation for each coupling step. After appropriate deblocking of 2, segments 2 and 3 were coupled by the DCC-HOBT method, followed by complete deblocking and ion-exchange chromatographic purification, affording the C-terminal half of thymosin alpha 1, H-Asp-Leu-Lys-Glu-Lys-Lys-Glu-Val-Val-Glu-Glu-Ala-Glu-Asn-OH (1).