title: Structure of the nisin leader peptidase NisP revealing a C-terminal autocleavage activity.
abstract: Nisin is a widely used antibacterial lantibiotic polypeptide produced by Lactococcus lactis. NisP belongs to the subtilase family and functions in the last step of nisin maturation as the leader-peptide peptidase. Deletion of the nisP gene in LAC71 results in the production of a non-active precursor peptide with the leader peptide unremoved. Here, the 1.1 Å resolution crystal structure of NisP is reported. The structure shows similarity to other subtilases, which can bind varying numbers of Ca atoms. However, no calcium was found in this NisP structure, and the predicted calcium-chelating residues were placed so as to not allow NisP to bind a calcium ion in this conformation. Interestingly, a short peptide corresponding to its own 635-647 sequence was found to bind to the active site of NisP. Biochemical assays and native mass-spectrometric analysis confirmed that NisP possesses an auto-cleavage site between residues Arg647 and Ser648. Further, it was shown that NisP mutated at the auto-cleavage site (R647P/S648P) had full catalytic activity for nisin leader-peptide cleavage, although the C-terminal region of NisP was no longer cleaved. Expressing this mutant in L. lactis LAC71 did not affect the production of nisin but did decrease the proliferation rate of the bacteria, suggesting the biological significance of the C-terminal auto-cleavage of NisP.
authors.affiliation: State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, People's Republic of China.
authors.forename: Yueyang
authors.initials: Y
authors.lastname: Xu
authors.affiliation: State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, People's Republic of China.
authors.forename: Xin
authors.initials: X
authors.lastname: Li
authors.affiliation: State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, People's Republic of China.
authors.forename: Ruiqing
authors.initials: R
authors.lastname: Li
authors.affiliation: State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, People's Republic of China.
authors.forename: Shanshan
authors.initials: S
authors.lastname: Li
authors.affiliation: State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, People's Republic of China.
authors.forename: Hongqian
authors.initials: H
authors.lastname: Ni
authors.affiliation: State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, People's Republic of China.
authors.forename: Hui
authors.initials: H
authors.lastname: Wang
authors.affiliation: State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, People's Republic of China.
authors.forename: Haijin
authors.initials: H
authors.lastname: Xu
authors.affiliation: State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, People's Republic of China.
authors.forename: Weihong
authors.initials: W
authors.lastname: Zhou
authors.affiliation: Department of Applied Chemistry and Microbiology, Division of Microbiology, University of Helsinki, Helsinki, Finland.
authors.forename: Per E J
authors.initials: PE
authors.lastname: Saris
authors.affiliation: State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, People's Republic of China.
authors.forename: Wen
authors.initials: W
authors.lastname: Yang
authors.affiliation: State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, People's Republic of China.
authors.forename: Mingqiang
authors.initials: M
authors.lastname: Qiao
authors.affiliation: State Key Laboratory of Medicinal Chemical Biology, College of Life Sciences, Nankai University, Tianjin, People's Republic of China.
authors.forename: Zihe
authors.initials: Z
authors.lastname: Rao
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry*; D018360:Crystallography, X-Ray; D013294:Lactococcus lactis / Q000737:chemistry*; D013058:Mass Spectrometry; D008565:Membrane Proteins / Q000737:chemistry*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011487:Protein Conformation; D059748:Proteolysis; D013381:Subtilisins / Q000737:chemistry*
chemical_list: D001426:Bacterial Proteins; D008565:Membrane Proteins; C080937:NisP protein, Lactococcus lactis; D013381:Subtilisins
keywords: NisP; nisin; nisin leader peptidase