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

Autor: Frank Oling, Peter H. Pouwels, Rudy A. Demel, Egbert Smit, Beatriz Martínez
Přispěvatelé: Electron Microscopy
Rok vydání: 2001
Předmět:
Models
Molecular
crystallization
domain
Protein Structure
Secondary
Lactobacillus acidophilus
Protein structure
Cell Wall
Sequence Analysis
Protein
Structural Biology
NUCLEOTIDE-SEQUENCE
Trypsin
Peptide sequence
Membrane Glycoproteins
biology
Lactobacillus crispatus
Proteolytic enzymes
food and beverages
REGULAR ARRAYS
Solutions
Biochemistry
CHEMICAL CHARACTERIZATION
Electrophoresis
Polyacrylamide Gel
S-layer
Protein Binding
EXPRESSION
Lactobacillus casei
Recombinant Fusion Proteins
Molecular Sequence Data
Phosphatidylserines
CLONING
Bacterial Proteins
Escherichia coli
Amino Acid Sequence
SURFACE-LAYERS
Protein Structure
Quaternary
Molecular Biology
Lactobacillus helveticus
STRAINS
Membrane Proteins
AEROMONAS-SALMONICIDA
biology.organism_classification
GENE
Molecular biology
Peptide Fragments
ATCC-4356
Protein Structure
Tertiary
Microscopy
Electron
cell wall binding
Sequence Alignment
S-layer protein
Zdroj: Journal of Molecular Biology, 305(2), 245-257. Academic Press
ISSN: 0022-2836
Popis: Lactobacillus acidophilus, like many other bacteria, harbors a surface layer consisting of a protein (SA-protein) of 43 kDa. SA-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 A; b = 53 A, and γ = 102°) resembling those determined for the S-layer of Lactobacillus helveticus ATCC 12046. Sequence comparison of SA-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 SA-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 SA-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 SA-protein. A calculated SA-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. © 2001 Academic Press. Chemicals/CAS: 1,2-dioleoylphosphatidylserine, 70614-14-1; Bacterial Proteins; Membrane Glycoproteins; Membrane Proteins; Peptide Fragments; Phosphatidylserines; Recombinant Fusion Proteins; Solutions; surface array protein, bacteria; Trypsin, EC 3.4.21.4
Databáze: OpenAIRE
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