Engineering unusual amino acids into peptides using lantibiotic synthetase

Jun Ichi Nagao, Kouki Shioya, Yoshitaka Harada, Ken Ichi Okuda, Takeshi Zendo, Jiro Nakayama, Kenji Sonomoto

Research output: Chapter in Book/Report/Conference proceedingChapter

9 Citations (Scopus)

Abstract

Alteration of protein structure and function by introducing unusual amino acids has great potential to develop new biological tool and to produce novel therapeutic agents. Lantibiotics produced by Gram-positive bacteria are ribosomally synthesized and post-translationally modified antimicrobial peptides. The modification enzyme involved in lantibiotic biosynthesis can catalyze the formation of unusual amino acids in the nascent lantibiotic prepeptide. Here, a novel methodology on the lantibiotic nukacin ISK-1 is described for engineering unusual amino acid residues into hexa-histidine-tagged (His-tagged) prepeptide NukA by the modification enzyme NukM in Escherichia coli. Co-expression of His-tagged NukA and NukM, purification of the resulting His-tagged prepeptide by affinity chromatography, and subsequent mass spectrometry analysis show that the prepeptide is converted into a postulated peptide with decrease in mass which results from the formation of unusual amino acids such as dehydrated amino acid, lanthionine, or 3-methyl lanthionine at the expected positions. The modified prepeptide can be readily obtained by one-step purification. This strategy will thus be a simple and powerful tool for introducing unusual amino acid residues aimed at peptide engineering.

Original languageEnglish
Title of host publicationHeterologous Gene Expression in E.coli
Subtitle of host publicationMethods and Protocols
EditorsThomas Evans, Jr., Ming-Qun Xu
Pages225-236
Number of pages12
DOIs
Publication statusPublished - Dec 1 2011

Publication series

NameMethods in Molecular Biology
Volume705
ISSN (Print)1064-3745

Fingerprint

Bacteriocins
Ligases
Amino Acids
Peptides
Histidine
Gram-Positive Bacteria
Enzymes
Affinity Chromatography
Mass Spectrometry
Escherichia coli
Proteins

All Science Journal Classification (ASJC) codes

  • Molecular Biology
  • Genetics
  • Medicine(all)

Cite this

Nagao, J. I., Shioya, K., Harada, Y., Okuda, K. I., Zendo, T., Nakayama, J., & Sonomoto, K. (2011). Engineering unusual amino acids into peptides using lantibiotic synthetase. In T. Evans, Jr., & M-Q. Xu (Eds.), Heterologous Gene Expression in E.coli: Methods and Protocols (pp. 225-236). (Methods in Molecular Biology; Vol. 705). https://doi.org/10.1007/978-1-61737-967-3_13

Engineering unusual amino acids into peptides using lantibiotic synthetase. / Nagao, Jun Ichi; Shioya, Kouki; Harada, Yoshitaka; Okuda, Ken Ichi; Zendo, Takeshi; Nakayama, Jiro; Sonomoto, Kenji.

Heterologous Gene Expression in E.coli: Methods and Protocols. ed. / Thomas Evans, Jr.; Ming-Qun Xu. 2011. p. 225-236 (Methods in Molecular Biology; Vol. 705).

Research output: Chapter in Book/Report/Conference proceedingChapter

Nagao, JI, Shioya, K, Harada, Y, Okuda, KI, Zendo, T, Nakayama, J & Sonomoto, K 2011, Engineering unusual amino acids into peptides using lantibiotic synthetase. in T Evans, Jr. & M-Q Xu (eds), Heterologous Gene Expression in E.coli: Methods and Protocols. Methods in Molecular Biology, vol. 705, pp. 225-236. https://doi.org/10.1007/978-1-61737-967-3_13
Nagao JI, Shioya K, Harada Y, Okuda KI, Zendo T, Nakayama J et al. Engineering unusual amino acids into peptides using lantibiotic synthetase. In Evans, Jr. T, Xu M-Q, editors, Heterologous Gene Expression in E.coli: Methods and Protocols. 2011. p. 225-236. (Methods in Molecular Biology). https://doi.org/10.1007/978-1-61737-967-3_13
Nagao, Jun Ichi ; Shioya, Kouki ; Harada, Yoshitaka ; Okuda, Ken Ichi ; Zendo, Takeshi ; Nakayama, Jiro ; Sonomoto, Kenji. / Engineering unusual amino acids into peptides using lantibiotic synthetase. Heterologous Gene Expression in E.coli: Methods and Protocols. editor / Thomas Evans, Jr. ; Ming-Qun Xu. 2011. pp. 225-236 (Methods in Molecular Biology).
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