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Applied and Environmental Microbiology, December 2008, p. 7613-7619, Vol. 74, No. 24
0099-2240/08/$08.00+0 doi:10.1128/AEM.00789-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.
Binding Specificity of the Lantibiotic-Binding Immunity Protein NukH
Ken-ichi Okuda,1
Sae Yanagihara,1
Kouki Shioya,1
Yoshitaka Harada,1
Jun-ichi Nagao,1
Yuji Aso,2
Takeshi Zendo,1
Jiro Nakayama,1 and
Kenji Sonomoto1,3*
Laboratory of Microbial Technology, Division of Microbial Science and Technology, Department of Bioscience and Biotechnology, Faculty of Agriculture, Graduate School, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan,1 Laboratory of Food Science, Department of Environmental Education for Human Life, Faculty of Education, Shimane University, 1060 Nishikawatsu, Matsue, Shimane 690-8504, Japan,2 Laboratory of Functional Food Design, Department of Functional Metabolic Design, Bio-Architecture Center, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan3
Received 7 April 2008/ Accepted 23 October 2008
NukH is a lantibiotic-binding immunity protein that shows strong binding activity against type A(II) lantibiotics. In this study, the binding specificity of NukH was analyzed by using derivatives of nukacin ISK-1, which is a type A(II) lantibiotic produced by Staphylococcus warneri ISK-1. Interactions between cells of Lactococcus lactis transformants expressing nukH and nukacin ISK-1 derivatives were analyzed by using a quantitative peptide-binding assay. Differences in the cell-binding rates of each nukacin ISK-1 derivative suggested that three lysine residues at positions 1 to 3 of nukacin ISK-1 contribute to the effective binding of nukacin ISK-1 to nukH-expressing cells. The binding levels of mutants with lanthionine and dehydrobutyrine substitutions (S11A nukacin4-27 and T24A nukacin4-27, respectively) to nukH-expressing cells were considerably lower than those of nukacin4-27, suggesting that unusual amino acids play a decisive role in NukH recognition. Additionally, it was suggested that T9A nukacin4-27, a mutant with a 3-methyllanthionine substitution, binds to NukH via an intermolecular disulfide bond after it is weakly recognized by NukH. We succeeded in the detection of specific type A(II) lantibiotics from the culture supernatants of various bacteriocin producers by using the binding specificity of nukH-expressing cells.
* Corresponding author. Mailing address: Laboratory of Microbial Technology, Division of Microbial Science and Technology, Department of Bioscience and Biotechnology, Faculty of Agriculture, Graduate School, Kyushu University, 6-10-1 Hakozaki, Higashi-ku, Fukuoka 812-8581, Japan. Phone and fax: 81-92-642-3019. E-mail: sonomoto{at}agr.kyushu-u.ac.jp
Published ahead of print on 31 October 2008.
Applied and Environmental Microbiology, December 2008, p. 7613-7619, Vol. 74, No. 24
0099-2240/08/$08.00+0 doi:10.1128/AEM.00789-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.
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