Automated high-throughput synthesis of artificial glycopeptides. Small-molecule probes for chemical glycobiology

A fully automated method for the synthesis of artificial glycopeptides having two (similar or different) carbon-linked glycosyl moieties on a dipeptide scaffold has been developed. By use of this approach that combines the diversity of peptide/pseudopeptide and glycosides, different glycoside moieties can be incorporated onto the peptide/pseudopeptide backbone in a highly controlled manner. The approach utilizes a stepwise reductive amination with glycoside aldehyde derivatives (model 1) or (ii) glycoside reductive amination followed by glycoside amide bond formation (model 2). Further, an automated method has been utilized in the high-throughput library synthesis of 4 x 96 artificial glycopeptides. These libraries were tested as chemical probes/inhibitors of enzyme systems that convert a glucose moiety into rhamnose prior to incorporation of the rhamnose unit and the conversion of UDP-galactopyranose to UDP-galactofuranose via UDP-galactopyranose mutase enzyme during the biosynthesis of the mycobacterium cell wall.     

An expedient synthesis of orthogonally protected lysinoalanine from Aloc-protected Garner’s aldehyde

An expedient synthesis of orthogonally protected lysinoalanine has been developed. We have prepared a novel Garner’s aldehyde derivative bearing an Aloc group; reductive amination of this aldehyde with Fmoc-Lys-OPMB gave the lysinoalanine skeleton. This was then transformed into an orthogonally protected lysinoalanine derivative suitable for the synthesis of side-chain bridged cyclic peptides by solid phase peptide synthesis methods.     

Combinatorial synthesis of RGD model cyclic peptides utilizing a palladium-catalyzed carbonylative macrolactamization on a polymer support

A combinatorial synthesis of 24-member RGD models was accomplished on polymer-support. Ortho-, meta-, and para-iodobenzylamines loaded on an aldehyde linker by reductive amination were coupled with RGD sequences and various omega-amino acids by a split-and-pool method. Palladium-catalyzed carbonylative macrolactamization of the polymer-supported cyclization precursors, followed by acid cleavage, provided conformationally restricted RGD model cyclic peptides.     

Characterization of aldehyde-PEG tethered surfaces: influence of PEG chain length on the specific biorecognition

A functionalized poly(ethylene glycol) (PEG) layer possessing a reactive aldehyde group at the free end of the tethered PEG chain was constructed by simple coating on the substrate, using the acetal-PEG/poly(DL-lactide) block copolymer, followed by the hydrolysis of the acetal end group by an acid treatment. The reactivity of the aldehyde group at the distal end of the PEG tethered chain was evaluated via a reductive amination using 4-amino-2,2,6,6-tetramethylpiperidinyloxy as the model compound. Further conjugation of the aldehyde group with sugar moieties has demonstrated an increased recognition ability with lectins with an increasing PEG chain length, which was attributable to the mobility of the chain end. These results provide a novel idea for highly sensitive biorecognition, suggesting a method to create highly selective biosensing surfaces that are able to prevent the undesired nonspecific adsorption of biocomponents.     

Design and synthesis of dimeric heparinoid mimetics

Synthetic oligosaccharide constructs exhibiting tailored and well-defined heparan sulfate (HS) like sequences offer the potential to modulate dynamic HS-dependent biomolecular recognition processes. We report an efficient strategy for the generation of HS-like fragments [GlcA-beta-(1,4)-GlcNAc] and related dimerized (gemini) disaccharides (4a and 4b) via n-pentenyl glycoside formation. When a convergent synthetic approach was utilized, construction of target molecules was achieved through a combination of chemoselective protection/deprotection protocols, imidate and n-pentenyl glycosylations, and functional group manipulations followed by ozonolysis and reductive amination. For example, glycosylation of a 2-azido glycoside (25) with a trichloroacetimidate glucuronic acid donor (13), using a catalytic amount of TMSOTf, furnished heparin-like disaccharides (28a and 28b) that were equipped with an n-pentenyl tether at the anomeric end. In turn, heparinoid-like gemini disaccharides (4a and 4b) were produced by selective transformation of the olefinic unit in the n-pentenyl glycoside to the four-carbon aldehyde followed by reductive amination with ethylenediamine. The described synthetic approach provides access to structural variants of small heparinoid oligomers as versatile building blocks for generating novel HS mimetic pharmacotherapeutics, diagnostic reagents, and biomaterials.     

N-Terminal peptide aldehydes as electrophiles in combinatorial solid phase synthesis of novel peptide isosteres

N-Terminal peptide aldehydes were synthesized on a solid support and utilized as electrophiles in nucleophilic reactions in order to furnish novel and diverse peptide isosteres. The aldehyde moiety of the peptide was synthesized by coupling a protected aldehyde building block to the peptide and deprotecting it quantitatively in less than 3 min. It was found that protection of the two succeeding amide nitrogens was necessary in order to avoid reaction between the aldehyde and backbone amides. The N-terminal peptide aldehydes were successfully reacted in the following way: (a) reductive amination with a large variety of amines, leading to N-alkyl-gamma-aminobutyric peptide isosteres positioned centrally in the peptide; (b) reductive amination with amino esters, leading to N-terminal 2,5-diketopiperazine peptides; (c) Horner-Wadsworth-Emmons olefination, leading to unsaturated peptide isosteres positioned centrally in the peptide; and (d) Pictet-Spengler condensations, leading to tetrahydro-beta-carbolines either positioned centrally in a peptide or fused with a diketopiperazine ring in the N-terminus of the peptide.     

Synthesis of the aldehyde of oligomeric polyoxyethylene

Conjugates of polyoxyethylene with proteins and surfaces are of much importance in various biotechnical applications. Consequently, methods for preparing the aldehyde of POE are of interest because of the potential ready attachment of this compound to proteins and aminated surfaces by reductive amination. In this work we describe the results of attempts to prepare an oligomeric POE aldehyde by six routes, two of which proved to be effective. Two previously published routes, manganese-oxide and DMSO/Ac₂O oxidation were ineffective. The aldehyde was shown to be unstable in the presence of base and thus of little use for reductive amination. On the other hand, the benzaldehyde derivative, which we also synthesized, is stable, easy to prepare, and undergoes ready reductive amination.     

A new route to 1,4-oxazepanes and 1,4-diazepanes from Garner aldehyde

A highly efficient and convenient method for the synthesis of substituted chiral 1,4-oxazepanes and 1,4-diazepanes have been described from Garner aldehyde through reductive amination with amino ester hydrochlorides followed by intramolecular cyclization as the key steps.     

Synthesis of N alpha-(1-phenyl-2-mercaptoethyl) amino acids, new building blocks for ligation and cyclization at non-cysteine sites: scope and limitations in peptide synthesis

A new and convenient method for the synthesis and incorporation of N(alpha)-(1-phenyl-2-mercaptoethyl)-derivatized amino acids applicable to chemical ligation at non-cysteine sites is presented. N(alpha)-Auxiliary derivatives of glycine and alanine were easily prepared using reductive amination approaches. Several strategies for the incorporation of these derivatives into peptide chains were investigated: coupling without protection, with acid-labile protection, with base-labile protection, and via a novel protection strategy using the thiazolidine derivative. All amino acid derivatives were successfully coupled to various peptide resins, and with the exception of those incorporating Boc-protected derivatives, all resins yielded the desired peptide fragments. However, the coupling of the two alanine derivative diastereomers generated some epimerization. Finally, N-terminal auxiliary glycine and alanine peptides were cyclized, and the corresponding native circular peptides were obtained upon successful removal of the auxiliary.     

Enantioselective total synthesis of Wieland-Gumlich aldehyde and (-)-strychnine

A total synthesis of (-)-strychnine in 15 steps from 1,3-cyclohexanedione in 0.15% overall yield is described. The sequence followed in the assembling of rings is: E-->AE [2-(2-nitrophenyl)-1,3-cyclohexanedione]-->ACE (3a-aryloctahydroindol-4-one)-->ACDE (arylazatricyclic core)-->ABCDE (strychnan skeleton)-->ABCDEF (Wieland-Gumlich aldehyde)-->ABCDEFG (strychnine). The key steps of the synthesis are the enantioselective construction of the 3a-(2-nitrophenyl)-octahydroindol-4-one ring system and the closure of the piperidine ring by a reductive Heck cyclization to generate the pivotal intermediate (-)-14. In contrast, a Lewis acid promoted a-alkoxypropargylic silane-enone cyclization did not lead to synthetically useful azatricyclic ACDE intermediates. The introduction of C-17 and the closure of the indoline ring by reductive amination of the alpha-(2-nitrophenyl) ketone moiety complete the strychnan skeleton from which, via the Wieland-Gumlich aldehyde, the synthesis of (-)-strychnine is achieved.     

Thiophene backbone amide linkers, a new class of easily prepared and highly acid-labile linkers for solid-phase synthesis

Solid-phase synthesis is of tremendous importance for small-molecule and biopolymer synthesis. Linkers (handles) that release amide-containing products after completion of solid-phase synthesis are widely used. Here we present a new class of highly acid-labile backbone amide linkers (BAL handles) based on 3,4-ethylenedioxythiophene (EDOT), which we have termed T-BAL. These thiophene linkers are synthesized in three convenient steps from commercially available EDOT. In the linker design, the spacer was introduced to the EDOT core either via a carbon-carbon bond or via a thioether linkage. Introduction of the spacer via a C-C bond was performed by a chemoselective Negishi coupling without transient protection of the aldehyde group to provide the T-BAL1 handle. Introduction via a thioether linkage was performed by a facile nucleophilic aromatic substitution between the brominated EDOT aldehyde and unprotected mercapto acids to provide T-BAL2 and T-BAL3 handles. The minimal use of protecting groups gave the corresponding linker molecules in few synthetic steps and in good yields. After anchoring of the linker to a polymeric support, introduction of the first amino acid was achieved by reductive amination, giving a secondary amine. A following acylation of the secondary amine with a symmetrical amino acid anhydride resulted in a backbone amide linkage between the handle and the growing substrate (e.g., peptide chain). After solid-phase synthesis, the substrates could be released from the resin by either low acid conditions using 1% TFA in CH2Cl2 or high acid conditions such as 50% TFA in CH2Cl2. Peptide thioesters could be released from the T-BAL1 handle under very mild conditions using aqueous acetic acid. Tert-butyl based protecting groups, tert-butyl esters, tert-butyl ethers, and Boc groups, as well as dimethyl acetals were relatively stable to these mild conditions for release of the peptides.     

Selective chemical modifications of dextran

Specific modification of the reducing end group of dextran has been achieved using the reductive amination procedure and solvent systems designed to optimize polymer reactivity. Dextran fraction (with M _w ranging from 10,000 to 500,000 daltons) were derivatized with [¹⁴C]-octadecylamine in yields of up to 60% to afford the corresponding alkyl dextrans which are of interest as affinity ligands. Reactive dextran intermediates with terminal amine, carboxyl, and aldehyde functions were prepared using sodium cyanoborohydride and ammonium acetate, glycine, and glucosamine, respectively. The dextran glucosamine derivative was further modified by reductive amination with octadecylamine. Similarly, condensation of dextran with streptomycin produced a new type of cationic derivative bearing a terminal, branched saccharide residue. Other reducing-end modifications included nitroxide-spin labelling, covalent attachment to aminopropyl-activated glass beads, and a carbodiimide-mediated amidation of carboxyl—dextran. The reductive amination method was also applied to guar gum and locust bean gum.     

Role of the peri-effect in synthesis and reactivity of highly substituted naphthaldehydes: a novel backbone amide linker for solid-phase synthesis

Handles (linkers) with an aldehyde functionality that permits the anchoring of substrates by reductive amination have, since their first report in the mid-1990s, become widely-used tools in solid-phase synthesis. In the synthesis of peptides, they allow anchoring of the growing peptide chain through a backbone amide, thus giving easy access to C-terminal modified or cyclic peptides. Recently, we described two new handles (NAL-1 and NAL-2) with dialkoxynaphthaldehyde core structures. Here, we describe the design, synthesis and properties of a novel trialkoxynaphthalene-based backbone amide linker (NAL-3). The NAL-3 handle is based on a trialkoxynaphthaldehyde (NALdehyde-3) that was synthesized in nine high-yielding steps from 3-methoxyphenylacetic acid in 51% overall yield. The naphthalene ring system was constructed using a regioselective methanesulfonic acid-catalyzed ring-closing reaction. The tetra-substituted naphthalene derivative 1,3,6-trimethoxynaphthalene-2-carbaldehyde (7) was selectively demethylated in the 1 position using BBr(3). The selectivity of this reaction is discussed, based on the crystal structures of reactant and product, 1-hydroxy-3,6-dimethoxy-naphthalene-2-carbaldehyde (8), and in the context of the peri-effect. The new handle was anchored to an aminomethylated poly(styrene) solid support, followed by assembly of a model dipeptide, then a study of the cleavage properties under acidic conditions was carried out. Surprisingly, the trialkoxynaphthaldehyde-based handle proved less acid-labile than the dialkoxynaphthaldehyde handles, and this fact is discussed with respect to handle design.     

Chemical Synthesis of beta-Homonojirimycin, of Its N-Butyl Derivative, and of "Methyl Homoazacellobioside"

beta-Homonojirimycin (2) was prepared by the highly stereoselective double reductive amination of a 2,6-heptodiulose derivative (6 or 13) using ammonium formate and NaBH(3)CN. The process was unsuccessful with primary amines. The synthesis of N-butyl-beta-homonojirimycin (19) was achieved by the N-butanoylation of a derivative of 2 followed by the reduction of the resulting tertiary amide. Compound 19 was found to be completely devoid of anti-HIV activity, in marked contrast with N-butyl-1-deoxynojirimycin. The coupling of the 1-O-p-toluenesulfonyl derivative of 2, compound 20, with methyl 2,3,6-tri-O-benzyl-alpha-D-glucopyranoside, followed by a deprotection step, provided pseudodisaccharide 23, the "homoaza" analog of methyl alpha-cellobioside and a potential inhibitor of beta-glucan-processing enzymes.     

A solid-phase synthetic strategy for labeled peptides: synthesis of a biotinylated derivative of the delta opioid receptor antagonist TIPP (Tyr-Tic-Phe-Phe-OH)

A general solid-phase synthetic strategy for labeled peptides was developed and used to prepare a biotinylated derivative of the delta opioid receptor antagonist TIPP (Tyr-Tic-Phe-Phe-OH). A monoprotected hydrophilic diamine linker was attached to an aldehyde-containing solid-phase resin by reductive amination, followed by introduction of biotin and peptide synthesis to yield Tyr-Tic-Phe-Phe-Asp-NH(CH(2)CH(2)O)(2)CH(2)CH(2)NH-biotin (2). The high delta receptor affinity and selectivity of 2 demonstrate the applicability of this design approach for labeled peptide derivatives.     

Synthesis of an immunomodulator (+)-conagenin and its analogs

Stereoselective synthesis of an immunomodulator (+)-conagenin was achieved. Both amine and carboxylic acid moieties were prepared from commercially available optically active methyl 3-hydroxy-2-methylpropanoate using dirhodium(II)-catalyzed C-H amination and chelation-controlled reductions as key steps. In addition, demethyl analogs of conagenin were synthesized using similar procedures.     

alpha-Ketocarbonyl peptides: a general approach to reactive resin-bound intermediates in the synthesis of peptide isosteres for protease inhibitor screening on solid support.

alpha-Ketocarbonyl peptides were generated from peptide precursors on solid support via a metal-ion-catalyzed transamination. The reaction proceeded to completion within 2 h with glyoxylate as electrophile and copper(II) ions as catalyst in an aqueous acetate buffer at pH 5.5-6.0. The variety of naturally occurring alpha-amino acid substrates gave rise to a diverse set of differentially functionalized ketones. The highly reactive terminal ketocarbonyls were prone to aldol-type dimerization and could be transferred into stable moieties by oxime formation, reduction to the alcohol, or reductive amination, respectively. The alpha-ketocarbonyl peptides were efficient in nucleophilic addition of C-nucleophiles such as phosphono-ylides and allylsilanes.     

Enzyme–Polymer Conjugates to Enhance Enzyme Shelf Life in a Liquid Detergent Formulation

Herein, the synthesis of enzyme–polymer conjugates is reported. Four different activated polymers (mPEG‐aldehyde, mPEG‐NHS, maltodextrin‐aldehyde, carboxymethyl cellulose aldehyde) are conjugated to the surface of protease, α‐amylase, and lipase using two different strategies (reductive amination and alkylation with NHS‐activated acid). Although the chemical modification of the enzymes is accompanied by losses in enzyme activity (maximum loss 40%), the covalent attachment of polymers increases the thermal stability and the stability in a standard detergent formulation compared to the unmodified enzymes. The enzyme–polymer conjugates are characterized by asymmetrical‐flow field‐flow fractionation and differential scanning microcalorimetry. Furthermore, it is demonstrated that conjugated enzymes still show performance in a real washing process. Enzyme–polymer conjugates show a potential as a stabilizing system for enzymes in detergents.     

Modular synthesis of pyrrolo[2,1‐b]thiazoles and related monocyclic pyrrolo structures

A modular synthesis of selectively‐substituted pyrrolo[2,1‐b]thiazoles (Δ⁶ isomeric form) has been implemented, involving a distinctive bicyclization reaction of a mucobromic acid derivative followed by a Suzuki‐Miyaura coupling. A novel process of Δ⁶ to Δ⁷ isomerization of the pyrrolothiazole structure was uncovered that appears to involve a 1,4‐addition‐1,2‐elimination mechanism. Preparation of 1,5‐dihydropyrrol‐2‐one structures, selectively substituted at the 3‐ and 4‐positions, was also achieved using the mucobromic acid synthon in a reductive amination process. J. Heterocyclic Chem., (2011).     

First Total Synthesis of Hyacinthacine A6 from the Protected Derivative of Polyhydroxylated Pyrrolidine

（1S、2R、3R、5R、7aR）-1,2-Dihydroxy-3-hydroxy methyl-5-methylpyrrolizidine（hyacinthacine A6, I） was synthesized by Wittig＇s methodology via the reaction of aldehyde 6, prepared from the partially protected derivative of polyhydroxylated pyrrolidine, with appropriated ylides, followed by cyclization through the intemal reductive amination process of the resulting a,B-unsaturated ketone 7, and total deprotection.