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Synthesis and application of dipeptides; current status and perspectives

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Abstract

The functions and applications of l-α-dipeptides (dipeptides) have been poorly studied compared with proteins or amino acids. Only a few dipeptides, such as aspartame (l-aspartyl-l-phenylalanine methyl ester) and l-alanyl-l-glutamine (Ala-Gln), are commercially used. This can be attributed to the lack of an efficient process for dipeptide production though various chemical or chemoenzymatic method have been reported. Recently, however, novel methods have arisen for dipeptide synthesis including a nonribosomal peptide-synthetase-based method and an l-amino acid α-ligase-based method, both of which enable dipeptides to be produced through fermentative processes. Since it has been revealed that some dipeptides have unique physiological functions, the progress in production methods will undoubtedly accelerate the applications of dipeptides in many fields. In this review, the functions and applications of dipeptides, mainly in commercial use, and methods for dipeptide production including already proven processes as well as newly developed ones are summarized. As aspartame and Ala-Gln are produced using different industrial processes, the manufacturing processes of these two dipeptides are compared to clarify the characteristics of each procedure.

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References

  • Abumard NN, Morse EL, Lochs H, Williams PE, Adibi SA (1989) Possible sources of glutamine for parenteral nutrition: impact on glutamine metabolism. Am J Physiol 257:E228–E234

    Google Scholar 

  • Aboulmagd E, Oppermann-Sanio FB, Steinbuchel A (2001) Purification of Synechocystis sp. strain PCC6308 cyanophycin synthetase and its characterization with respect to substrate and primer specificity. Appl Environ Microbiol 67:2176–2182

    CAS  PubMed  PubMed Central  Google Scholar 

  • Adibi SA (1997) The oligopeptide transporter (Pept-1) in human intestine: biology and function. Gastroenterology 113:332–340

    CAS  PubMed  Google Scholar 

  • Ager DJ, Pantaleone DP, Henderson SA, Katritzky AR, Prakash I, Walters DE (1998) Commercial, synthetic nonnutritive sweeteners. Angew Chem Int Ed 37:1802–1817

    CAS  Google Scholar 

  • Albers S, Wernerman J, Stehle P, Vinnars E, Furst P (1988) Availability of amino acids supplied intravenously in healthy man as synthetic dipeptides: kinetic evaluation of L-alanyl-L-glutamine and glycyl-L-tyrosine. Clinical Sci 75:463–468

    CAS  Google Scholar 

  • Albini N, Auricchio S, Minisci F (1985) Base catalysis and solvent effect in the synthesis of aspartame. Chem Ind 15:484–485

    Google Scholar 

  • Ariyoshi Y, Nagao M, Naotake (1974a) Method of producing α-L-aspartyl-L-phenylalanine lower alkyl ester. US Patent no. 3,786,039

  • Ariyoshi Y, Yamatani T, Uchiyama N, Yasuda N, Toi K (1974b) Method of producing α-L-aspartyl-L-phenylalanine alkyl ester. US Patent no. US 3,833,553

  • Ashiuchi A, Misono H (2002) Biochemistry and molecular genetics of poly-γ-glutamate synthesis. Appl Microbiol Biotechnol 59:9–14

    CAS  PubMed  Google Scholar 

  • Babizhayev MA, Deyev AI, Yermakova VN, Semiletov YA, Davydova NG, Kurysheva NI, Zhukotskii AV, Goldman IM (2001) N-Acetylcarnosine, a natural histidine-containing dipeptide, as a potent ophthalmic drug in treatment of human cataracts. Peptides 22:979–994

    CAS  PubMed  Google Scholar 

  • Bachman GL, Oftedahl ML, Vineyard BD (1976) Process for the preparation of α-L-aspartyl-L-phenylalanine alkyl esters. US Patent no. US 3,933,781

  • Begum G, Cunliffe A, Leveritt M (2005) Physiological role of carnosine in contracting muscle. Int J Sport Nutr Exerc Metab 15:493–514

    CAS  PubMed  Google Scholar 

  • Belshaw PJ, Walsh CT, Stachelhaus T (1999) Aminoacyl-CoAs as probes of condensation domain selectivity in nonribosomal peptide synthesis. Science 284:486–489

    CAS  PubMed  Google Scholar 

  • Bergmann M, Fraenkel-Conrat H (1937) The role of specificity in the enzymatic synthesis of proteins. J Biol Chem 119:707–720

    CAS  Google Scholar 

  • Bergmann M, Fraenkel-Conrat H (1938) The enzymatic synthesis of peptide bonds. J Biol Chem 124:1–6

    CAS  Google Scholar 

  • Bongers J, Heimer EP (1994) Recent applications of enzymatic peptide synthesis. Peptides 15:183–193

    CAS  PubMed  Google Scholar 

  • Bordusa F (2002) Proteases in organic synthesis. Chem Rev 102:4817–4867

    CAS  PubMed  Google Scholar 

  • Candela T, Fouet A (2006) Poly-gamma-glutamate in bacteria. Mol Microbiol 60:1091–1098

    CAS  PubMed  Google Scholar 

  • Cho CH, Luk CT, Ogle CW (1991) The membrane-stabilizing action of zinc carnosine (Z-103) in stress-induced gastric ulceration in rats. Life Sci 49:PL189–PL194

    CAS  PubMed  Google Scholar 

  • Cloninger MR, Baldwin RE (1970) Aspartyl phenylalanine methyl ester: a low-calorie sweetener. Science 170:81–82

    CAS  PubMed  Google Scholar 

  • de Armas RR, Diaz HG, Molina R, Gonzalez MP, Uriarte E (2004) Stochastic-based descriptors studying peptides biological properties: modeling the bitter tasting threshold of dipeptides. Bioorg Med Chem 12:4815–4822

    Google Scholar 

  • de Ferra F, Rodriguez F, Tortora O, Tosi C, Grandi G (1997) Engineering of peptide synthetases. J Biol Chem 272:25304–25309

    PubMed  Google Scholar 

  • Dieckmann R, Neuhof T, Pavela-Vrancic M, von Dohren H (2001) Dipeptide synthesis by an isolated adenylate-forming domain of non-ribosomal peptide synthetase (NRPS). FEBS Lett 498:42–45

    CAS  PubMed  Google Scholar 

  • Doekel S, Marahiel MA (2000) Dipeptide formation on engineered hybrid peptide synthetases. Chem Biol 7:373–384

    CAS  PubMed  Google Scholar 

  • Doel MT, Eaton M, Cook EA, Lewis H, Patel T, Carey NH (1980) The expression in E. coli of synthetic repeating polymeric genes coding for poly(L-asapartyl-L-phenylalanine). Nucleic Acids Res 8:4575–4592

    CAS  PubMed  PubMed Central  Google Scholar 

  • Duerfahrt T, Doekel S, Sonke T, Quaedflieg PJLM, Marahiel MA (2003) Construction of hybrid peptide synthetases for the production of α-L-aspartyl-L-phenylalanine, a precursor for the high-intensity sweetener aspartame. Eur J Biochem 270:4555–4563

    CAS  PubMed  Google Scholar 

  • Finking R, Marahiel MA (2004) Biosynthesis of nonribosomal peptides. Annu Rev Microbiol 58:453–487

    CAS  Google Scholar 

  • Francois P, Francis D, Pierre M (1990) Enzyme, its method of production and its application to the preparation of methyl N-(L-aspartyl-1) L-phenylananinate. US Patent no. US 4,916,062

  • Furst P (2001) New developments in glutamine delivery. J. Nutr 131(9 suppl):2562S–2568S

    CAS  PubMed  Google Scholar 

  • Furst P, Pfaender P, Werner F (1985) Glutaminhaltige Aminosaure-Zubereltungen. EP Patent no. EP 0087750

  • Furst P, Pogan K, Stehle P (1997) Glutamine dipeptides in clinical nutrition. Nutrition 13:731–737

    CAS  PubMed  Google Scholar 

  • Galperin MY, Koonin EV (1997) A diverse superfamily of enzymes with ATP-dependent carboxylate-amine/thiol ligase activity. Protein Sci 6:2639–2643

    CAS  PubMed  PubMed Central  Google Scholar 

  • Goeters C, Wenn A, Mertes N, Wempe C, Van Aken H, Stehle P, Bone H-G (2002) Parenteral L-alanyl-L-glutamine improves 6-month outcome in critically ill patients. Crit Care Med 30:2032–2037

    CAS  PubMed  Google Scholar 

  • Gulewitsch W, Amiradzibi S (1900) Ueber das Carnosin, eine neue organishe Base des Fleishextractes. Ber Deutsch Chem Ges 33:1902–1903

    CAS  Google Scholar 

  • Guiotto A, Calderan A, Ruzza P, Borin G (2005) Carnosine and carnosine-related antioxidants: a review. Curr Med Chem 12:2293–2315

    CAS  PubMed  Google Scholar 

  • Hashimoto S (2006) Occurrence, biosynthesis, and biotechnological production of dipeptides. Microbiol Monogr 5:327–348

    Google Scholar 

  • Henriques V, Gjaldbak IK (1911) Untersuchungen uber die Plasteinbildung. Z Physiol Chem 71:485–517

    CAS  Google Scholar 

  • Hill JB, Gelman Y, Dryden, Jr HL, Erickson R, Hsu K, Johnson MR (1991) One-pot process for the preparation of α-L-aspartyl-L-phenylalanine methyl ester hydrochloride. US Patent no. US 5,053,532

  • Hines HM, Sutfin DC (1956) Physiologic properties of anserine and carnosine. Am J Physiol 186:286–288

    CAS  PubMed  Google Scholar 

  • Ikeda H, Yagasaki M, Hashimoto S (2006) Methods for manufacturing dipeptides or their derivatives. WO Patent application no. 2006/001382

  • Inouye K, Kusano M, Hashida Y, Minoda M, Yasukawa K (2007) Engineering, expression, purification, and production of recombinant thermolysin. Biotechnol Annu Rev 13:43–64

    CAS  PubMed  Google Scholar 

  • Isowa Y, Ohmori M, Ichikawa T, Mori K, Nonaka Y, Kihara K, Oyama K, Satoh H, Nishimura S (1979) The thermolysi-catalyzed condensation reactions of n-substituted aspartic and glutamic acids with phenylalanine alkyl esters. Tetrahedron Lett 20:2611–2612

    Google Scholar 

  • Katsoyannis PG, Ginos JZ (1969) Chemical synthesis of peptides. Annu Rev Biochem 38:881–912

    CAS  PubMed  Google Scholar 

  • Kayser H, Meisel H (1996) Stimulation of human peripheral blood lymphocytes by bioactive peptides derived from bovine milk proteins. FEBS Lett 383:18–20

    CAS  PubMed  Google Scholar 

  • Keller U, Schauwecker F (2003) Combinatorial biosynthesis of non-ribosomal peptides. Comb Chem High Throug Scre 6:527–540

    CAS  Google Scholar 

  • Khavinson VK, Anisimov VN (2000) Synthetic dipeptide vilon (L-Lys-L-Glu) increases life span and inhibits a development of spontaneous tumors in mice. Doklady Akad Nauk 372:421–423

    CAS  Google Scholar 

  • Kino K, Nakazawa Y, Yagasaki M (2006) Method for producing dipeptide. WO Patent application no. 2006/101023

  • Kino K, Nakazawa Y, Yagasaki M (2007) Method for production of dipeptide. WO Patent application no. 2007/074858

  • Kino K, Kotanaka Y, Yagasaki M (2008) Method for production of dipeptide. WO Patent application no. 2008/038613

  • Kitts DD, Weiler K (2003) Bioactive proteins and peptides from food sources. Applications of bioprocesses used in isolation and recovery. Curr Pharm Design 9:1309–1323

    CAS  Google Scholar 

  • Kumar D, Bhalla TC (2005) Microbial proteases in peptide synthesis: approaches and applications. Appl Microbiol Biotechnol 68:726–736

    CAS  PubMed  Google Scholar 

  • Linne U, Marahiel MA (2004) Reactions catalyzed by mature and recombinant nonribosomal peptide synthetases. Methods in Enzymol 388:293–315

    CAS  Google Scholar 

  • Lombard C, Saulnier J, Wallach JM (2005) Recent trends in protease-catalyzed peptide synthesis. Protein Peptide Lett 12:621–629

    CAS  Google Scholar 

  • Matsufuji H, Matsui T, Seki E, Osajima K, Nakashima M, Osajima Y (1994) Angiotensin I-converting enzyme inhibitory peptides in an alkaline protease hydrolyzate derived from sardine muscle. Biosci Biotech Biochem 58:2244–2245

    CAS  Google Scholar 

  • Meister A (1974) Glutathione synthesis. The Enzyme 10:671–697

    CAS  Google Scholar 

  • Mootz HD, Kessler N, Linne U, Eppelmann K, Schwarzer D, Marahiel MA (2002) Decreasing the ring size of a cyclic nonribosomal peptide antibiotic by in-frame module deletion in the biosynthetic genes. J Am Chem Soc 124:10980–10981

    CAS  PubMed  Google Scholar 

  • Morihara K (1987) Using proteases in peptide synthesis. TIBTECH 5:164–170

    CAS  Google Scholar 

  • Nakanishi K, Kamikubo T, Matsuno R (1985) Continuous synthesis of N-(benzyloxycarbonyl)-L-aspartyl-L-phenylalanine methyl ester with immobilized thermolysin in an organic solvent. Bio/Technol 3:459–464

    CAS  Google Scholar 

  • Nakanishi K, Takeuchi A, Matsuno R (1990) Long-term continuous synthesis of aspartame precursor in a column reactor with an immobilized thermolysin. Appl Microbiol Biotechnol 32:633–636

    CAS  Google Scholar 

  • Nilsson BL, Soellner MB, Raines RT (2005) Chemical synthesis of proteins. Annu Rev Biophys Biomol Struct 34:91–118

    CAS  PubMed  PubMed Central  Google Scholar 

  • Nitta A, Nishioka H, Fukumitsu H, Furukawa Y, Sugiura H, Shen L, Furukawa S (2004) Hydrophobic dipeptide Leu-Ile protects against neuronal death by inducing brain-derived neurotrophic factor and glial cell line-derived neurotrophic factor synthesis. J Neurosci Res 78:250–258

    CAS  PubMed  Google Scholar 

  • Oyama K, Irino S, Hagi N (1987) Production of aspartame by immobilized thermoase. Methods in Enzymol 136:503–516

    CAS  Google Scholar 

  • Rausch C, Weber T, Kohlbacher O, Wohleben W, Huson DH (2005) Specificity prediction of adenylation domains in nonribosomal peptide synthetases (NRPS) using transductive support vector machines (TSVMs). Nucleic Acids Res 33:5799–5808

    CAS  PubMed  PubMed Central  Google Scholar 

  • Roth E, Ollenschlager G, Hamilton G, Simmel A, Langer K, Fekl W, Jakesz R (1988) Influence of two glutamine-containing dipeptides on growth of mammalian cells. In Vitro Cell Dev Biol 24:696–698

    CAS  PubMed  Google Scholar 

  • Sano T, Sugaya T, Inoue K, Mizutani S, Ono Y, Kasai M (2000) Process research and development of L-alanyl-L-glutamine, a component of parenteral nutrition. Org Process Res Dev 4:147–152

    CAS  Google Scholar 

  • Sato M, Hosokawa T, Yamaguchi T, Nakano T, Muramoto K, Kahara T, Funayama K, Kobayashi A, Nakano T (2002) Angiotensin I-converting enzyme inhibitory peptides derived from wakame (Undaria pinnatifida) and their antihypertensive effect in spontaneously hypertensive rats. J Agric Food Chem 50:6245–6252

    CAS  PubMed  Google Scholar 

  • Schellenberger V, Jakubke HD (1991) Protease-catalyzed kinetically controlled peptide synthesis. Angew Chem Int Ed Engl 30:1437–1449

    Google Scholar 

  • Schiffman SS (1976) Taste of dipeptides. Physiol Behavior 17:523–535

    CAS  Google Scholar 

  • Sieber SA, Marahiel MA (2005) Molecular mechanisms underlying nonribosomal peptide synthesis: approaches to new antibiotics. Chem Rev 105:715–738

    CAS  PubMed  Google Scholar 

  • Sinisterra JV, Alcantara AR (1993) Synthesis of peptides catalyzed by enzymes: a practical overview. J Mol Catalysis 84:327–364

    CAS  Google Scholar 

  • Stachelhaus T, Schneider A, Marahiel MA (1995) Rational design of peptide antibiotics by targeted replacement of bacterial and fungal domains. Science 269:69–72

    CAS  PubMed  Google Scholar 

  • Stachelhaus T, Mootz HD, Marahiel MA (1999) The specificity-conferring code of adenylation domains in nonribosomal peptide synthetases. Chem Biol 6:493–505

    CAS  PubMed  Google Scholar 

  • Stehle P, Pfaender P, Furst P (1984) Isotachophoretic analysis of a synthetic dipeptide L-alanyl-L-glutamine. Evidence for stability during heat sterilization. J Chromatogr 294:507–512

    CAS  Google Scholar 

  • Suzuki T, Hirano T, Suyama M (1987) Free imidazole compounds in white and dark muscles of migratory marine fish. Comp Biochem Physiol B 87:615–619

    CAS  PubMed  Google Scholar 

  • Symmank H, Franke P, Saenger W, Bernhard F (2002) Modification of biologically active peptides: production of a novel lipohexapeptide after engineering of Bacillus subtilis surfactin synthetase. Protein Engine 15:913–921

    CAS  Google Scholar 

  • Tabata K, Hashimoto S (2005) Microorganisms producing dipeptides and process for producing dipeptides using the microorganisms. WO Patent application no. 2005/045006

  • Tabata K, Hashimoto S (2007) Fermentative production of L-alanyl-L-glutamine by a metabolically engineered Escherichia coli strain expressing L-amino acid α-ligase. Appl Environ Microbiol 73:6378–6385

    CAS  PubMed  PubMed Central  Google Scholar 

  • Tabata K, Ikeda H, Hashimoto S (2005) ywfE in Bacillus subtilis codes for a novel enzyme, L-amino acid ligase. J Bacteriol 187:5195–5202

    CAS  PubMed  PubMed Central  Google Scholar 

  • Takagi H, Shimoi H, Ueda H, Amano H (1979) Morphine-like analgesia by a new dipeptide, L-tyrosyl-L-arginine (kyotorphin) and its analogue. Eur J Pharmacol 55:109–111

    CAS  PubMed  Google Scholar 

  • Walsh CT (1989) Enzymes in the D-alanine branch of bacterial cell wall peptidoglycan assembly. J Biol Chem 264:2393–2396

    CAS  PubMed  Google Scholar 

  • Ye L, Ramstrom O, Ansell RJ, Masson M-O, Masbach K (1999) Use of molecularly imprinted polymers in a biotransformation process. Biotechnol Bioengi 64:650–655

    CAS  Google Scholar 

  • Yokoyama K, Chiba H, Yoshikawa M (1992) Peptide inhibitors for angiotensin I-converting enzyme from thermolysin digest of dried bonito. Biosci Biotechnol Biochem 56:1541–1545

    CAS  PubMed  Google Scholar 

  • Yokozeki K, Hara S (2005) A novel and efficient enzymatic method for the production of peptides from unprotected starting materials. J Biotechnol 115:211–220

    CAS  PubMed  Google Scholar 

  • Yukawa T, Kawasaki T, Nakamura M, Yamashita T, Tuji T (1994) JP Patent application JP Patent no. JP 06/80075

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  1. Technical Research Laboratories of Kyowa Hakko Kogyo Co., Ltd., 1-1 Kyowa-cho, Hofu, 747-8522, Japan

    Makoto Yagasaki & Shin-ichi Hashimoto

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  1. Makoto Yagasaki
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  2. Shin-ichi Hashimoto
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Correspondence to Makoto Yagasaki.

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Yagasaki, M., Hashimoto, Si. Synthesis and application of dipeptides; current status and perspectives. Appl Microbiol Biotechnol 81, 13–22 (2008). https://doi.org/10.1007/s00253-008-1590-3

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