The S-layer Protein of Lactobacillus acidophilus ATCC 4356: Identification and Characterisation of Domains Responsible for S-protein Assembly and Cell Wall Binding

doi:10.1006/jmbi.2000.4258

Journal of Molecular Biology

Volume 305, Issue 2, 12 January 2001, Pages 245-257

https://doi.org/10.1006/jmbi.2000.4258 Get rights and content

Abstract

Lactobacillus acidophilus, like many other bacteria, harbors a surface layer consisting of a protein (S_A-protein) of 43 kDa. S_A-protein could be readily extracted and crystallized in vitro into large crystalline patches on lipid monolayers with a net negative charge but not on lipids with a net neutral charge. Reconstruction of the S-layer from crystals grown on dioleoylphosphatidylserine indicated an oblique lattice with unit cell dimensions (a=118 Å; b=53 Å, and γ=102 °) resembling those determined for the S-layer of Lactobacillus helveticus ATCC 12046. Sequence comparison of S_A-protein with S-proteins from L. helveticus, Lactobacillus crispatus and the S-proteins encoded by the silent S-protein genes from L. acidophilus and L. crispatus suggested the presence of two domains, one comprising the N-terminal two-thirds (SAN), and another made up of the C-terminal one-third (SAC) of S_A-protein. The sequence of the N-terminal domains is variable, while that of the C-terminal domain is highly conserved in the S-proteins of these organisms and contains a tandem repeat. Proteolytic digestion of S_A-protein showed that SAN was protease-resistant, suggesting a compact structure. SAC was rapidly degraded by proteases and therefore probably has a more accessible structure. DNA sequences encoding SAN or Green Fluorescent Protein fused to SAC (GFP-SAC) were efficiently expressed in Escherichia coli. Purified SAN could crystallize into mono and multi-layered crystals with the same lattice parameters as those found for authentic S_A-protein. A calculated S_A-protein minus SAN density-difference map revealed the probable location, in projection, of the SAC domain, which is missing from the truncated SAN peptide. The GFP-SAC fusion product was shown to bind to the surface of L. acidophilus, L. helveticus and L. crispatus cells from which the S-layer had been removed, but not to non-stripped cells or to Lactobacillus casei.

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^☆: Abbreviations used: S, crystalline surface; SLH, S-layer homology region; DS, dialyzate supernatant; DP, dialyzate pellet

^☆☆: Edited by W. Baumeister

^f1: Present address: B. Martinez, Instituto de Productos Lacteos de Asturias, CSIC, Apdo. 85, 33300 Villaviciosa, Spain.

^f2: Corresponding author

^f3: E-mail address of the corresponding author: [email protected]

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Journal of Molecular Biology

Regular ArticleThe S-layer Protein of Lactobacillus acidophilus ATCC 4356: Identification and Characterisation of Domains Responsible for S-protein Assembly and Cell Wall Binding☆,☆☆

Abstract

Curr. Opin. Struct. Biol.

J. Biol. Chem.

J. Struct. Biol.

J. Biol. Chem.

FEMS Microbiol. Letters

Methods Enzymol.

J. Mol. Biol.

J. Mol. Biol.

J. Ultrastruct. Res.

J. Mol. Biol.

Principles of organization in eubacterial and archaebacterial surface proteins

Canad. J. Microbiol.

A rapid alkaline extraction procedure for screening recombinant plasmid DNA

Nucl. Acids Res.

The surface layer protein genes of Lactobacillus acidophilus, PhD thesis

Expression, secretion and antigenic variation of bacterial S-layer proteins

Mol. Microbiol.

S-layer protein of Lactobacillus acidophilus ATCC 4356: purification, expression in Escherichia coli, and nucleotide sequence of the corresponding gene

J. Bacteriol.

Identification, cloning and nucleotide sequence of a silent S-layer protein gene of Lactobacillus acidophilus ATCC 4356 which has extensive similarity with the S-layer protein gene of this species

J. Bacteriol.

The presence of two S-layer protein encoding genes is conserved among species related to Lactobacillusacidophilus

Microbiology

Cell wall of Thermoanaerobacterium thermosulfurigenes EM1: isolation of its components and attachment of the xylanase XynA

Arch. Microbiol.

The S-layer gene of Lactobacillus helveticus CNRZ 892: cloning, sequence and heterologous expression

Microbiology

The S-layer protein of Corynebacterium glutamicum is anchored to the cell wall by its C-terminal hydrophobic domain

Mol. Microbiol.

Monomolecular layers in the study of biomembranes

Subcell. Biochem.

The S-layer proteins of two Bacillus stearothermophilus wild-type strains are bound via their N-terminal region to a secondary cell wall polymer of identical chemical composition

J. Bacteriol.

Evidence that an N-terminal S layer protein fragment triggers the release of a cell-associated high-molecular-weight amylase in Bacillus stearothermophilus ATCC 12980

J. Bacteriol.

Regular Article
The S-layer Protein of Lactobacillus acidophilus ATCC 4356: Identification and Characterisation of Domains Responsible for S-protein Assembly and Cell Wall Binding☆,☆☆