Synthesis of OmpA Protein of Escherichia coli K12 in Bacillus subtilis

Kallio, Pauli T.; Simonen, Mika; Palva, Ilkka; Sarvas, Matti

doi:10.1099/00221287-132-3-677

Cited by 14 publications

(15 citation statements)

References 56 publications

(27 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…After centrifugation (5000 x g, 10 min at 4°C) the pellets were resuspended In 50 mM TrIs-HCl, pH 8. The specific rabbit antisera were anti-OmpA sera KH497 (the immunogen was the W-terminal 24 kDa trypsin fragment of OmpA protein of E. coli obtained by trypsin treatment of envelope fraction followed by purification in preparative SDS-PAGE as described by Kallio et al (1986)) and KH686 (the immunogen was the Bac-OmpA228OmpA228-ss treated with trypsin which resulted in formation of two trypsin fragments of about 24 kDa and 28 kDa), and anti-OmpF serum KH526 (OmpF protein used in immunization was purified by preparative SDS-PAGE (Sarvas and Nurminen, 1985) from E. co//K-12 EH277 (derivative of strain CE1215 containing plasmid pJP33 described in Tommassen et ai, 1982)) and anti-TEM-[i-lactamase serum KD12-4 (the rabbits were immunized with TEM p-lactamase purified from E. coli carrying the run-off plasmid pKN410; Ehrlich, 1978).…”

Section: Other Methodsmentioning

confidence: 99%

Secretion of the Escherichia coli outer membrane proteins OmpA and OmpF in Bacillus subtilis is blocked at an early intracellular step

Puohiniemi

Simonen

Muttilainen

et al. 1992

Molecular Microbiology

View full text Add to dashboard Cite

When the genes coding for the outer membrane (OM) proteins OmpA and OmpF of Escherichia coli are fused to a signal sequence of a bacillar exoenzyme and expressed in Bacillus subtilis they remain cell-bound and the signal sequence is not cleaved. To identify the step of arrest in the export of these proteins we studied their accessibility to protease applied to intact protoplasts; they remained resistant indicating fully intracellular localization. Both proteins appeared associated with the cell membranes in sedimentation and flotation centrifugation experiments. However, OmpA and OmpF proteins synthesized in B. subtilis without a signal sequence were similarly associated with membranes in centrifugation experiments whereas electron microscopy showed the presence of intracytoplasmic inclusion bodies not obviously attached to the cytoplasmic membrane. We conclude that OmpA and OmpF proteins even when provided with a functional signal sequence do not enter the export pathway in B. subtilis, probably owing to lack of a specific export component in B. subtilis.

show abstract

Section: Other Methodsmentioning

confidence: 99%

Secretion of the Escherichia coli outer membrane proteins OmpA and OmpF in Bacillus subtilis is blocked at an early intracellular step

Puohiniemi

Simonen

Muttilainen

et al. 1992

Molecular Microbiology

View full text Add to dashboard Cite

show abstract

“…Two separate methods were used to prepare OmpA. Firstly, the TX envelopes (5 mg of protein) were treated with trypsin for 2 h at 37°C and electrophoresed in 15% preparative SDS-acrylamide gel (Kallio et al, 1986). The OmpA of mol-wt 24 x lo3 was eluted from the gel with SDS 0.05% at 37°C overnight.…”

Section: Bacteriamentioning

confidence: 99%

Protective capacity of antibodies to outer-membrane components of Escherichia coli in a systemic mouse peritonitis model

Vuopio‐Varkila

Karvonen²,

Saxén³

1988

Journal of Medical Microbiology

View full text Add to dashboard Cite

“…OmpA, when fused to the signal peptide of B. amyloliquefaciens a-amylase, was not translocated across the cytoplasmic membrane of B. subtilis. Kallio et al (1986) suggested that this may be due to the absence in B. subtilis of a specific factor required for the translocation of OmpA across the membrane.…”

Section: Discussionmentioning

confidence: 99%

Non-functional expression of Escherichia coli signal peptidase I in Bacillus subtilis

Dijl¹,

Jong²,

Smith³

et al. 1991

Journal of General Microbiology

View full text Add to dashboard Cite

The Escherichia cofi fep gene, encoding signal peptidase I (SPase I) was provided with Baciffus subtifis transcription/translation signals and expressed in this organism. When present on a low-copy-number plasmid, the amount of E. cofi SPase I produced (per mg cell protein) in B. subtifis was half that produced in wild-type E. cofi cells. The production of E. cofi SPase I in B. subtifis was increased approximately fivefold by cloning the fep gene into a high-copy-number plasmid. The expression of E cofi SPase I in B. subtifis did not appear to increase the rate of processing of two hybrid secretory precursor proteins. Two observations may explain the failure of E. cofi SPase I to stimulate processing of exported proteins in B. subtifis. First, the E. cofiSPase I was apparently not exposed on the outside of the B. subtifis cytoplasmic membrane, indicating its incorrect insertion into the membrane. Second, in vi?ro processing studies, using cell-free extracts of B. subtilis producing E. cofi SPase I, suggested that the enzyme was not active. A further outcome of this study was that conditions favouring processing of precursors by SPase I in cell-free extracts of E. cofi did not favour processing by the corresponding enzyme in B. subtifis cell-free extracts. This suggests that significant differences exist between the two enzymes. The observation that antibodies directed against E. cofi SPase I did not cross-react with B. subtifis membrane proteins supports this idea.

show abstract

Synthesis of OmpA Protein of Escherichia coli K12 in Bacillus subtilis

Cited by 14 publications

References 56 publications

Secretion of the Escherichia coli outer membrane proteins OmpA and OmpF in Bacillus subtilis is blocked at an early intracellular step

Secretion of the Escherichia coli outer membrane proteins OmpA and OmpF in Bacillus subtilis is blocked at an early intracellular step

Protective capacity of antibodies to outer-membrane components of Escherichia coli in a systemic mouse peritonitis model

Non-functional expression of Escherichia coli signal peptidase I in Bacillus subtilis

Contact Info

Product

Resources

About