Direct preparation of peptide thioesters using an Fmoc solid-phase method

Li, Xiangqun; Kawakami, Toru; Aimoto, Saburo

doi:10.1016/s0040-4039(98)01868-1

Cited by 198 publications

(129 citation statements)

References 16 publications

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“…strain ADP1acr1⍀Km, on May 12, 2018 by guest http://aem.asm.org/ but major amounts of dihexadecyl disulfides were formed, leading to a decrease of the acyl acceptor concentration and thus most probably lowering the thio wax ester yields. Thioesters are used as activated esters (12,17,24) for the synthesis of peptides, for macrocyclic antibiotics, and for various other pharmaceuticals by organic chemists and, in particular, the pharmaceutical industry (4,7,11,14,16,19). If the amounts of thio wax ester accumulated by Acinetobacter sp.…”

Section: Discussionmentioning

confidence: 99%

“…In the present study, we therefore investigated whether alkanethiols like 1-hexadecanthiol, 1,8-octanedithiol, and 1-S-monopalmitoyloctanedithiol can serve as alternative acyl acceptors in the WS/DGAT-mediated acyl transfer reaction, resulting in the biosynthesis of unusual thio wax esters. Thioesters are used as activated esters (12,17,24) for the synthesis of peptides, macrocyclic antibiotics, and various other pharmaceuticals in organic chemistry and, in particular, in the pharmaceutical industry (4,7,11,14,16,19).…”

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confidence: 99%

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Thio Wax Ester Biosynthesis Utilizing the Unspecific Bifunctional Wax Ester Synthase/Acyl Coenzyme A:Diacylglycerol Acyltransferase of Acinetobacter sp. Strain ADP1

Uthoff

Stöveken

Weber

et al. 2005

Appl Environ Microbiol

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The bifunctional wax ester synthase/acyl coenzyme A (acyl-CoA):diacylglycerol acyltransferase (WS/DGAT) from Acinetobacter sp. strain ADP1 (formerly Acinetobacter calcoaceticus ADP1) mediating the biosyntheses of wax esters and triacylglycerols was used for the in vivo and in vitro biosynthesis of thio wax esters and dithio wax esters. For in vitro biosynthesis, 5His 6 WS/DGAT comprising an N-terminal His 6 tag was purified from the soluble protein fraction of Escherichia coli Rosetta(DE3)pLysS (pET23a::5His 6 atf). By employing SP-Sepharose high-pressure and Ni-nitrilotriacetic acid fast-protein liquid chromatographies, a 19-fold enrichment with a final specific activity of 165.2 nmol mg of protein ؊1 min ؊1 was achieved by using 1-hexadecanol and palmitoyl-CoA as substrates. Incubation of purified 5His 6 WS/DGAT with 1-hexadecanethiol and palmitoylCoA as substrates resulted in the formation of palmitic acid hexadecyl thio ester (10.4% relative specific activity of a 1-hexadecanol control). Utilization of 1,8-octanedithiol and palmitoyl-CoA as substrates led to the formation of 1-S-monopalmitoyloctanedithiol and minor amounts of 1,8-S-dipalmitoyloctanedithiol (59.3% relative specific activity of a 1-hexadecanol control). The latter dithio wax ester was efficiently produced when 1-S-monopalmitoyloctanedithiol and palmitoyl-CoA were used as substrates (13.4% specific activity relative to that of a 1-hexadecanol control). For the in vivo biosynthesis of thio wax esters, the knockout mutant Acinetobacter sp. strain ADP1acr1⍀Km, which is unable to produce fatty alcohols, was used. Cultivation of Acinetobacter sp. strain ADP1acr1⍀Km in the presence of gluconate, 1-hexadecanethiol, and oleic acid in nitrogen-limited mineral salts medium resulted in the accumulation of unusual thio wax esters that accounted for around 1.19% (wt/wt) of the cellular dry weight and consisted mainly of oleic acid hexadecyl thioester as revealed by gas chromatography-mass spectrometry.Polyhydroxyalkanoic acids (PHA) in bacteria represent the most abundant group of neutral storage lipids, which serve as intracellular carbon and energy storage compounds (23). In addition, triacylglycerols (TAGs) and wax esters also occur as bacterial storage lipids, though at a much lower frequency than in eukaryotic microorganisms (22). Significant TAG accumulation was found, particularly in species belonging to the class Actinomycetes (1), as was reported for the genera Mycobacterium (3), Nocardia, and Streptomyces (2). TAGs are synthesized by the acyl coenzyme A (acyl-CoA):diacylglycerol acyltransferase (DGAT) (13), which catalyzes the esterification of diacylglycerol with long-chain acyl-CoA. Wax esters are oxoesters of primary long-chain fatty alcohols and long-chain fatty acids. Occurrence of wax esters has been frequently reported for various Acinetobacter species (6). Under growthlimiting conditions, Acinetobacter calcoaceticus accumulates wax esters intracellularly as insoluble inclusions. The chemical structure of the wax esters synthesized by A. ca...

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Thio Wax Ester Biosynthesis Utilizing the Unspecific Bifunctional Wax Ester Synthase/Acyl Coenzyme A:Diacylglycerol Acyltransferase of Acinetobacter sp. Strain ADP1

Uthoff

Stöveken

Weber

et al. 2005

Appl Environ Microbiol

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show abstract

“…53,54 Applied for the first time for SPPS by two different laboratories independently. 55,56 Since then, several optimized removal conditions for SPS have been described, the most relevant being: 20% piperidine in DMF, 55 which is the most common, 1-5% DBU in DMF, 57,58 morpholine-DMF (1:1) 59 or 2% HOBt, 2% hexamethyleneimine, 25% N-methylpyrrolidine in DMSO-NMP (1:1), 60 Fmoc α-amino protection has been used for the SPS of several relevant peptides using the so-called Fmoc/ t Bu strategy, the production in Tm scale of the T20 peptide being one of the most important examples. 63 Nevertheless, the low solubility of some Fmoc derivatives in the most commonly used solvents for SPPS has stimulated the search for new base-labile protecting groups.…”

Section: Protecting Groups Removed By Basementioning

confidence: 99%

Amino Acid-Protecting Groups

2009

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“…Using similar strategy, Kajihara et al recently prepared N-terminal glycosylated peptide thioester of chemokine MCP-3 (27)(Scheme 2), and applied it for the total synthesis of MCP-3. Aimoto et al realized the direct synthesis of peptide thioester using the less nucleophilic base cocktail for the Fmoc removal (28). Using this method, we synthesized emmprin active domain carrying N-linked core pentasaccharide (29).…”

Section: Methodsmentioning

confidence: 99%