Proteases in Organic Synthesis

Bordusa, Frank

doi:10.1021/cr010164d

Cited by 272 publications

(163 citation statements)

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“…The concentrations of acyl donors (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15)(16), acyl acceptors (a-c), and enzymes were 1 mM, 20 mM, and 5 mM, respectively. The progress of the peptide coupling reaction was monitored by HPLC under the following conditions: isocratic elution at 1 ml/min, 0.1% trifluoroacetic acid/acetonitrile.…”

Section: Methodsmentioning

confidence: 99%

“…Especially, peptide synthesis using protease-catalyzed reverse reaction has been extensively studied with a variety of amino acids and peptide derivatives as coupling components. [2][3][4][5][6][7][8] It has been reported that the protease-catalyzed peptide synthesis is superior to the chemical coupling method because it is highly stereoselective, racemization-free, and requires less side-chain protection. The major drawback of the enzymatic method, however, is the respective substrate specificity.…”

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confidence: 99%

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Atlantic Cod Trypsin-Catalyzed Peptide Synthesis with Inverse Substrates as Acyl Donor Components

Fuchise

Kishimura

Zhi-yong

et al. 2010

CHEMICAL & PHARMACEUTICAL BULLETIN

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A large number of biologically active peptides have been isolated recently from bacterial, fungal, plant and animal sources and characterized in some detail. In particular, shortsequence peptides play important roles in the sensory appreciation of food toward four basic taste sensations (sweet, bitter, sour and salty). 1) Such peptides sometimes contain Damino acid and other unusual amino acids. Synthetic chemistry has witnessed remarkable progress with the development of novel biologically active peptides. Enzymatic peptide synthesis has emerged as a powerful approach to the preparation of short sequences. Especially, peptide synthesis using protease-catalyzed reverse reaction has been extensively studied with a variety of amino acids and peptide derivatives as coupling components. [2][3][4][5][6][7][8] It has been reported that the protease-catalyzed peptide synthesis is superior to the chemical coupling method because it is highly stereoselective, racemization-free, and requires less side-chain protection. The major drawback of the enzymatic method, however, is the respective substrate specificity. Thus, the application of proteases for peptide synthesis has not been fully investigated synthetic possiblity of a number of biologically significant peptides containing D-amino acid or other unusual amino acids. A few previous reports have shown the enzymatic condensation of noncorded amino acids, 9,10) peptide mimetics, 11) peptide conjugates, 12) and D-amino acid. 13) Previously, we reported that the inverse substrates such as p-amidino-14) and p-guanidinophenyl esters [15][16][17] behave as specific substrates for trypsin and trypsin-like enzymes and allow the specific introduction of an acyl group carrying a non-specific residue into the enzymatic active site. The characteristics of inverse substrates suggested that they are useful for enzymatic peptide synthesis. [18][19][20][21][22][23][24] Many studies on the characterization of trypsins from cold-adapted species have been reported. 25) These trypsins display substantially higher catalytic efficiency than their mammalian counterparts. [25][26][27][28][29][30][31][32][33] We previously reported the chum salmon trypsin-catalyzed synthesis of peptides using inverse substrates. 23,24) We had obtained commercially available Atlantic cod trypsin at almost the same time. We are interested in comparing of the catalytic efficiency of Atlantic cod trypsin with chum salmon trypsin. In the present work, we investigated Atlantic cod trypsin-catalyzed peptide synthesis using p-amidino-and p-guanidinophenyl esters of N atert-butyloxycarbonyl (Boc)-amino acid as acyl donor components with two types of acyl acceptors. We have found that Atlantic cod trypsin-catalyzed peptide synthesis of the Damino acid and b-amino acid-containing products is more efficient than chum salmon trypsin-catalyzed synthesis. Results and DiscussionThe kinetic constants for the trypsin-catalyzed hydrolysis of synthetic inverse substrates were analyzed on the basis of the following scheme.Where: Eϭe...

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Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

Atlantic Cod Trypsin-Catalyzed Peptide Synthesis with Inverse Substrates as Acyl Donor Components

Fuchise

Kishimura

Zhi-yong

et al. 2010

CHEMICAL & PHARMACEUTICAL BULLETIN

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“…The mutation of proteases is a powerful strategy to substantially change specific enzymatic properties, such as stability and substrate specificity. [28] Several proteases that have been engineered by site-directed or random mutagenesis have been studied for their aminolysis activity. [84,85] However, polymerization using these engineered proteases has not been previously examined.…”

Section: Modification Of Proteases For Polypeptide Synthesismentioning

confidence: 99%

“…The reverse reaction, aminolysis, can be used to create new amide bonds using appropriate amino acid substrates under suitable reaction conditions. [28,29] Hence, the tandem aminolysis of amino acid esters using proteases leads to polypeptide formation. This chemoenzymatic polymerization by proteases has several advantages compared to conventional condensation techniques.…”

Section: Introductionmentioning

confidence: 99%

Chemoenzymatic Synthesis of Polypeptides for Use as Functional and Structural Materials

Tsuchiya

Numata

2017

Macromolecular Bioscience

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Polypeptides inspired by the natural functional and structural proteins present in living systems are promising materials for various fields in terms of their versatile functionality and physical properties. Designing and synthesizing mimetic sequences of specific peptide motifs in proteins are important for exploring the functionality of natural proteins. Chemoenzymatic polymerization, which utilizes aminolysis (i.e., the reverse reaction of hydrolysis catalyzed by proteases), is a useful technique for synthesizing artificial polypeptide materials and has several advantages, including facile synthesis protocols, environmental friendliness, scalability, and atom economy. In this review, recent progress in chemoenzymatic polypeptide synthesis for the production of functional and structural materials for various applications is summarized in conjunction with the current status of technical challenges in the field.

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“…1 Proteases that also present esterase activity have been widely used to catalyze reactions related to peptide synthesis, such as ester hydrolysis, esterification, transesterification and peptide bond formation. 2,3 However, it is well known that the main concern in protease-catalyzed peptide bond synthesis is protease-catalyzed product consumption. [4][5][6] Therefore, the following approaches have been used to minimize such an undesirable reaction: (i) continuous removal of the product from the reaction medium by precipitation 7,8 or extraction; 9 (ii) freezing of the reaction; 10 (iii) use of an esterified N-acyl-amino acid or N α -acyl-peptide as acyl donor; 11 (iv) use of a mimetic substrate; 12 and (v) use of a chemically modified or mutant protease.…”

Section: Introductionmentioning

confidence: 99%

Dipeptide synthesis in biphasic medium: evaluating the use of commercial porcine pancreatic lipase preparations and the involvement of contaminant proteases

Liria

Romagna

Rodovalho

et al. 2008

J. Braz. Chem. Soc.

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Sínteses bem sucedidas de dipeptídeos a partir de Ac-L-Tyr-OEt ou Z-L-X-OMe (X: Asp, Tyr, Phe, Arg, Lys ou Thr) e glicina amidada em meio reacional bifásico foram realizadas usando dois tipos de preparações comerciais de lipase pancreática suína (PPL) (bruta (cPPL) e purificada (pPPL)). Nas mesmas condições reacionais, a α-quimotripsina, protease pancreática que também apresenta atividade esterásica, catalisou a síntese de Ac-L-Tyr-Gly-NH 2 com alta produtividade. Na maioria das sínteses também ocorreu a hidrólise do produto formado. Eletroforese em gel de poliacrilamida, ensaios enzimáticos com substratos cromogênicos específicos e cromatografia de exclusão molecular demonstraram que cPPL e pPPL apresentam proteases contaminantes e, portanto, atividades esterásica e amidásica. Em conjunto, esses resultados indicam que tais proteases, e não a PPL, possam ser os catalisadores majoritários da síntese da ligação peptídica quando ésteres de N α -acil-L-aminoácidos e preparações comerciais de PPL são usados. Por outro lado, tais dados não contradizem a possibilidade de usar cPPL como fonte barata de catalisadores para a síntese de dipeptídeos em condições reacionais brandas.Dipeptide syntheses starting from Ac-L-Tyr-OEt or Z-L-X-OMe (X: Asp, Tyr, Phe, Arg, Lys or Thr) and glycine amide in biphasic reaction media were achieved using two commercially available porcine pancreatic lipase (PPL) preparations (crude (cPPL) and purified PPL (pPPL)). Under the mild conditions employed, α-chymotrypsin, a pancreatic protease that also presents esterase activity, catalyzed Ac-L-Tyr-Gly-NH 2 synthesis with high productivity. Product hydrolysis also occurred in most of the syntheses studied. Polyacrylamide gel electrophoresis, enzymatic assays employing specific chromogenic substrates and size-exclusion chromatography revealed that cPPL and pPPL contain contaminant proteases and, therefore, exhibit esterase and amidase activities. Overall, these data indicate that those contaminants may be the main catalysts of peptide bond synthesis when N α -blocked-L-amino acid esters and the commercial PPL preparations are used. On the other hand, such data do not contest the possibility of using such enzyme preparations as an inexpensive source of catalysts for dipeptide synthesis under soft conditions. Keywords: biocatalysis, enzymes, peptide bond synthesis, n-hexane IntroductionIn the last decade, interest in environment friendly technologies has increased drastically. As a result, the ability of enzymes to catalyze reactions required for the manufacture of fine chemicals has been explored extensively. 1 Proteases that also present esterase activity have been widely used to catalyze reactions related to peptide synthesis, such as ester hydrolysis, esterification, transesterification and peptide bond formation. 2,3 However, it is well known that the main concern in protease-catalyzed peptide bond synthesis is protease-catalyzed product consumption. [4][5][6] Therefore, the following approaches have been used to minimize such an undesirable r...

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Proteases in Organic Synthesis

Cited by 272 publications

References 535 publications

Atlantic Cod Trypsin-Catalyzed Peptide Synthesis with Inverse Substrates as Acyl Donor Components

Atlantic Cod Trypsin-Catalyzed Peptide Synthesis with Inverse Substrates as Acyl Donor Components

Chemoenzymatic Synthesis of Polypeptides for Use as Functional and Structural Materials

Dipeptide synthesis in biphasic medium: evaluating the use of commercial porcine pancreatic lipase preparations and the involvement of contaminant proteases

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