S-layer protein of Lactobacillus acidophilus ATCC 4356: purification, expression in Escherichia coli, and nucleotide sequence of the corresponding gene.
The cell surfaces of several Lactobacillus species are covered by a regular layer composed of a single species of protein, the S-protein. The 43-kDa S-protein of the neotype strain Lactobacillus acidophilus ATCC 4356, which originated from the pharynx of a human, was purified. Antibodies generated against purified S-protein were used to screen a lambda library containing chromosomal L. acidophilus ATCC 4356 DNA. Several phages showing expression of this S-protein in Escherichia coli were isolated. A 4.0-kb DNA fragment of one of those phages hybridized to a probe derived from an internal tryptic fragment of the S-protein. The slpA gene, coding for the surface layer protein, was located entirely on the 4.0-kb fragment as shown by deletion analysis. The nucleotide sequence of the slpA gene was determined and appeared to encode a protein of 444 amino acids. The first 24 amino acids resembled a putative secretion signal, giving rise to a mature S-protein of 420 amino acids (44.2 kDa). The predicted isoelectric point of 9.4 is remarkably high for an S-protein but is in agreement with the data obtained during purification. The expression of the entire S-protein or of large, C-terminally truncated S-proteins is unstable in E. coli.
Boot
H J
HJ
TNO Medical Biological Laboratory, Rijswijk, The Netherlands.
Kolen
C P
CP
van Noort
J M
JM
Pouwels
P H
PH
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies; D001425:Bacterial Outer Membrane Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D001426:Bacterial Proteins; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial / Q000302:isolation & purification; D004926:Escherichia coli / Q000378:metabolism*; D005798:Genes, Bacterial; D007779:Lactobacillus acidophilus / Q000235:genetics / Q000378:metabolism*; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; D051379:Mice; D008807:Mice, Inbred BALB C / Q000276:immunology; D008969:Molecular Sequence Data; D015183:Restriction Mapping; D017386:Sequence Homology, Amino Acid
D000906:Antibodies; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; C055764:S-layer proteins