Skip to main content
Log in

Purification and characterization of Stenotrophomonas maltophilia-derived l-amino acid ester hydrolase for synthesizing dipeptide, isoleucyl-tryptophan

  • Original Article
  • Published:
3 Biotech Aims and scope Submit manuscript

Abstract

In the present study, we purified α-amino acid ester hydrolase (AEH) from cell-free extracts of the Stenotrophomonas maltophilia strain HS1. The approximately 70-kDa AEH from S. maltophilia HS1 (SmAEH) was homogeneous in sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) analyses, and was present as a tetramer in gel-filtration experiments. The activity of the SmAEH enzyme was then determined by monitoring the synthesis of the antihypertensive agent dipeptide isoleucyl-tryptophan (Ile-Trp) from isoleucyl methyl ester (Ile-OMe) and tryptophan (Trp). In these experiments, SmAEH had wide substrate specificity for acyl donors, such as Gly-OMe, β-Ala-OMe, Pro-OMe and Trp-OMe and Ile-OMe, and maximal activity were observed under conditions of pH 9.0 and 30 °C. SmAEH also showed the greatest stability at pH 9.0, whereas its activity was reduced by 40% after 10-min incubation at approximately 50 °C. In subsequent activity assays in the presence of various metal ions, Ag+ strongly inhibited enzyme activity. Finally, SmAEH activity was completely inhibited by phenylmethanesulfonyl fluoride (PMSF), suggesting that the protein is a serine protease.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  • Abe I, Hara S, Yokozeki K (2011) Gene cloning and characterization of α-amino acid ester acyl transferase in Empedobacter brevis ATCC14234 and Sphingobacterium siyangensis AJ2458. Biosci Biotechnol Biochem 75:2087–2092

    Article  CAS  Google Scholar 

  • Barends TR, Polderman-Tijmes JJ, Jekel PA, Hensgens CM, de Vries EJ, Janssen DB, Dijkstra BW (2003) The sequence and crystal structure of the α-amino acid ester hydrolase from Xanthomonas citri define a new family of β-lactam antibiotic acylases. J Biol Chem 278:23076–23084

    Article  CAS  Google Scholar 

  • Edgar RC (2004) MUSCLE: multiple sequence alignment with high accuracy and high throughput. Nucleic Acids Res 32:1792–1797

    Article  CAS  Google Scholar 

  • Enari H, Takahashi Y, Kawarasaki M, Tada M, Tatsuta K (2008) Identification of angiotensin I converting enzyme inhibitory peptides derived from salmon muscle and their antihypertensive effect. Fish Sci 74:911–920

    Article  CAS  Google Scholar 

  • Felsenstein J (1989) PHYLIP-phylogeny inference package (version 3.2). Cladistics 5:164–166

    Google Scholar 

  • Gill I, Vulfson EN (1993) Enzymic synthesis of short peptides in heterogeneous mixtures of substrates. J Am Chem Soc 115:3348–3349

    Article  CAS  Google Scholar 

  • Gill I, López-Fandiño R, Jorba X, Vulfson EN (1996) Biologically active peptides and enzymatic approaches to their production. Enzyme Microb Technol 18:162–183

    Article  CAS  Google Scholar 

  • Guzmán F, Barberis S, Illanes A (2007) Peptide synthesis: chemical or enzymatic. Electron J Biotechnol 10:279–314

    Article  Google Scholar 

  • Hansler M, Jakubke HD (1996) Nonconventional protease catalysis in frozen aqueous solutions. J Pept Sci 2:279–289

    Article  CAS  Google Scholar 

  • Hirao Y, Mihara Y, Kira I, Abe I, Yokozeki K (2013) Enzymatic production of l-alanyl-l-glutamine by recombinant E. coli expressing α-amino acid ester acyltransferase from Sphingobacterium siyangensis. Biosci Biotechnol Biochem 77:618–623

    Article  CAS  Google Scholar 

  • Iroyukifujita H, Eiichiyokoyama K, Yoshikawa M (2000) Classification and antihypertensive activity of angiotensin I-converting enzyme inhibitory peptides derived from food proteins. J Food Sci 65:564–569

    Article  Google Scholar 

  • Kato K (1980) Kinetics of acyl transfer by α-amino acid ester hydrolase from Xanthomonas citri. Agric Biol Chem 44:1083–1088

    CAS  Google Scholar 

  • Kato K, Kawahara K, Takahashi T, Igarasi S (1980) Enzymatic synthesis of amoxicillin by the cell-bound α-amino acid ester hydrolase of Xanthomonas citri. Agric Biol Chem 44:821–825

    CAS  Google Scholar 

  • Kim DJ, Byun SM (1990) Purification and properties of ampicillin acylase from Pseudomonas melanogenum. Biochimica et Biophysica Acta (BBA) Protein Struct Mol Enzymol 1040:12–18

    Article  CAS  Google Scholar 

  • Kino K, Nakazawa Y, Yagasaki M (2008a) Dipeptide synthesis by l-amino acid ligase from Ralstonia solanacearum. Biochem Biophys Res Commun 371:536–540

    Article  CAS  Google Scholar 

  • Kino K, Noguchi A, Nakazawa Y, Yagasaki M (2008b) A novel l-amino acid ligase from Bacillus licheniformis. J Biosci Bioeng 106:313–315

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  • Kurochkina V, Sklyarenko A, Berezina O, Yarotskii S (2013) Alpha-amino acid ester hydrolases: properties and applications. Appl Biochem Microbiol 49:672–694

    Article  CAS  Google Scholar 

  • Li GH, Le GW, Shi YH, Shrestha S (2004) Angiotensin I-converting enzyme inhibitory peptides derived from food proteins and their physiological and pharmacological effects. Nutr Res 24:469–486

    Article  CAS  Google Scholar 

  • Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275

    CAS  Google Scholar 

  • Malathi S, Chakraborty R (1991) Production of alkaline protease by a new Aspergillus flavus isolate under solid-substrate fermentation conditions for use as a depilation agent. Appl Environ Microbiol 57:712–716

    CAS  Google Scholar 

  • Mine Y, Li-Chan E, Jiang B (2011) Bioactive proteins and peptides as functional foods and nutraceuticals, vol 29. Wiley, New York

    Google Scholar 

  • Murray BA, Fitz Gerald RJ (2007) Angiotensin converting enzyme inhibitory peptides derived from food proteins: biochemistry, bioactivity and production. Curr Pharm Des 13:773–791

    Article  CAS  Google Scholar 

  • Ollis DL, Cheah Eong, Cygler Miroslaw, Dijkstra Bauke, Frolow Felix, Franken Sybille M, Harel Michal (1992) The α/β hydrolase fold. Protein Eng 5:197–211

    Article  CAS  Google Scholar 

  • Ono S, Hosokawa M, Miyashita K, Takahashi K (2006) Inhibition properties of dipeptides from salmon muscle hydrolysate on angiotensin I-converting enzyme. Int J Food Sci Technol 4:383–386

    Article  Google Scholar 

  • Polderman-Tijmes JJ et al (2002) Cloning, sequence analysis, and expression in Escherichia coli of the gene encoding an α-amino acid ester hydrolase from Acetobacter turbidans. Appl Environ Microbiol 68:211–218

    Article  CAS  Google Scholar 

  • Ryu YW, Ryu DD (1988) Semisynthetic β-lactam antibiotic synthesizing enzyme from Acetobacter turbidans: catalytic properties. Enzyme Microb Technol 10:239–245

    Article  CAS  Google Scholar 

  • Santos S, Torcato I, Castanho MA (2012) Biomedical applications of dipeptides and tripeptides. Pept Sci 98:288–293

    Article  CAS  Google Scholar 

  • Sato M et al (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

    Article  CAS  Google Scholar 

  • Sheldon RA, van Rantwijk F, van Langen LM, Wegman MA, Cao L, Janssen MHA (2001) Biocatalysts and biocatalysis in the synthesis of ß-Lactam antibiotics. In: Bruggink A (ed) Synthesis of β-Lactam antibiotics. Springer, Dordrecht, Netherlands, pp 102–148

  • Sugihara A, Shimada Y, Sugihara S, Nagao T, Watanabe Y, Tominaga Y (2001) A novel α-amino-acid esterase from Bacillus mycoides capable of forming peptides of DD-and DL-configurations. J Biochem 130:119–126

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  • Takahashi T, Yamazaki Y, Kato K (1974) Substrate specificity of an α-amino acid ester hydrolase produced by Acetobacter turbidans ATCC 9325. Biochem J 137:497–503

    Article  CAS  Google Scholar 

  • Yagasaki M, Hashimoto S (2008) Synthesis and application of dipeptides; current status and perspectives. Appl Microbiol Biotechnol 81:13–22

    Article  CAS  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

    Article  CAS  Google Scholar 

  • Yokozeki K, Suzuki S (2010) Method for producing dipeptides. US Patent 7,754,466. Ajinomoto Co Inc, Japan

Download references

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Mamoru Wakayama.

Ethics declarations

Conflict of interest

The authors declare that there is no conflict of interests.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplementary material 1 (DOCX 232 kb)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Hossain, M.S., Tanaka, T., Takagi, K. et al. Purification and characterization of Stenotrophomonas maltophilia-derived l-amino acid ester hydrolase for synthesizing dipeptide, isoleucyl-tryptophan. 3 Biotech 8, 173 (2018). https://doi.org/10.1007/s13205-018-1195-1

Download citation

  • Received:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1007/s13205-018-1195-1

Keywords

Navigation