title: Characterization of a S-layer protein from Lactobacillus crispatus K313 and the domains responsible for binding to cell wall and adherence to collagen.
abstract: It was previously shown that the surface (S)-layer proteins covering the cell surface of Lactobacillus crispatus K313 were involved in the adherence of this strain to human intestinal cell line HT-29. To further elucidate the structures and functions of S-layers, three putative S-layer protein genes (slpA, slpB, and slpC) of L. crispatus K313 were amplified by PCR, sequenced, and characterized in detail. Quantitative real-time PCR analysis reveals that slpA was silent under the tested conditions; whereas slpB and slpC, the putative amino acid sequences which exhibited minor similarities to the previously reported S-layer proteins in L. crispatus, were actively expressed. slpB, which was predominantly expressed in L. crispatus K313, was further investigated for its functional domains. Genetic truncation of the untranslated leader sequence (UTLS) of slpB results in a reduction in protein production, indicating that the UTLS contributed to the efficient S-layer protein expression. By producing a set of N- and C-terminally truncated recombinant SlpB proteins in Escherichia coli, the cell wall-binding region was mapped to the C terminus, where rSlpB(380-501) was sufficient for binding to isolated cell wall fragments. Moreover, the binding ability of the C terminus was variable among the Lactobacillus species (S-layer- and non-S-layer-producing strains), and teichoic acid may be acting as the receptor of SlpB. To determine the adhesion region of SlpB to extracellular matrix proteins, ELISA was performed. Binding to immobilized types I and IV collagen was observed with the His-SlpB(1-379) peptides, suggesting that the extracellular matrix protein-binding domain was located in the N terminus.
authors.affiliation: State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, People's Republic of China.
authors.forename: Zhilan
authors.initials: Z
authors.lastname: Sun
authors.forename: Jian
authors.initials: J
authors.lastname: Kong
authors.forename: Shumin
authors.initials: S
authors.lastname: Hu
authors.forename: Wentao
authors.initials: W
authors.lastname: Kong
authors.forename: Wenwei
authors.initials: W
authors.lastname: Lu
authors.forename: Wei
authors.initials: W
authors.lastname: Liu
mesh_terms: D018829:Adhesins, Bacterial / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics; D001665:Binding Sites; D002473:Cell Wall / Q000378:metabolism*; D003094:Collagen / Q000378:metabolism*; D004252:DNA Mutational Analysis; D004269:DNA, Bacterial / Q000737:chemistry / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D015870:Gene Expression; D020869:Gene Expression Profiling; D006801:Humans; D007778:Lactobacillus / Q000235:genetics* / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000235:genetics* / Q000378:metabolism*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011485:Protein Binding; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D017384:Sequence Deletion
chemical_list: D018829:Adhesins, Bacterial; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D008562:Membrane Glycoproteins; C055764:S-layer proteins; D003094:Collagen