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WO1994008011A1 - Proteine mutante recombinee derivee de pasteurella multocida et son procede de preparation - Google Patents

Proteine mutante recombinee derivee de pasteurella multocida et son procede de preparation Download PDF

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Publication number
WO1994008011A1
WO1994008011A1 PCT/DK1993/000312 DK9300312W WO9408011A1 WO 1994008011 A1 WO1994008011 A1 WO 1994008011A1 DK 9300312 W DK9300312 W DK 9300312W WO 9408011 A1 WO9408011 A1 WO 9408011A1
Authority
WO
WIPO (PCT)
Prior art keywords
protein
dna sequence
codon
codons
amino acid
Prior art date
Application number
PCT/DK1993/000312
Other languages
English (en)
Inventor
Svend Petersen
Ole C. Hansen
Holger K. Riemann
Lars Bjarne Nielsen
Original Assignee
Bioteknologisk Institut
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Bioteknologisk Institut filed Critical Bioteknologisk Institut
Priority to AU51081/93A priority Critical patent/AU5108193A/en
Publication of WO1994008011A1 publication Critical patent/WO1994008011A1/fr

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Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/195—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria
    • C07K14/285—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from bacteria from Pasteurellaceae (F), e.g. Haemophilus influenza
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies

Definitions

  • TECHNICAL BACKGROUND AND PRIOR ART PAR is a widespread, highly contagious infectious disease in animals, in particular pigs where the infection affects the normal bone structure of the snout as a result of nasal bone resorption causing deformation of the snout, sneezing, nasal discharge and bleeding from the nasal mucosa.
  • the disease causes substantial economic losses due to growth retardation and increased susceptibility to other infectious diseases.
  • rmPMT protein which was constructed, is one having four amino acid substitutions and designated
  • pSPE1234, pSPE1020 and pSPE1134 were deposited on 9 September 1993 with the DSM-Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH under the accession numbers DSM 8547, DSM 8548 and DSM 8546, respectively in the form of cultures of E. coli DH5 ⁇ transformed with these plasmids, except pSPE1134 which was deposited in E. coli K12 strain MC1000.
  • the codon (s) being substituted may be a codon or codons coding for an amino acid/amino acids selected from serine, histidine, glutamine and threonine and the substituting codon (s) may be a codon or codons coding for an amino acid or amino acids coding for valine, leucine or tyrosine. It is contemplated that useful rmPMTs may be proucked by having a hydrophilic amino acid replaced by a hydrophobic amino acid.
  • the mutagenization treatment of step (ii) may be selected so as to cause deletion of at least one codon coding for an amino acid of position between position 1131 and 1285.
  • the deletion(s) concern(s) at least one codon coding for an amino acid of position between position 1175 and 1285 such as e.g. deletion of at least one codon coding for an amino acid of position between position 1215 and 1285.
  • a plasmid may be a convenient replicon.
  • the selection of a suitable plasmid may be directed by well-known considerations, including the capability of plasmids to replicate in a wide range of host organisms, or the ability to occur in a host cell in a high copy number.
  • a particularly useful type of plasmids may be plasmids showing "runaway" replication behaviour. Examples of plasmids which in accordance with the invention may be useful, include a plasmid selected from pSPE680, pSPE888, pSPE900, pSPE1003, pSPE1020, pSPE1038, pSPE1134 and pSPE 1234.
  • the rmPMT-encoding gene may furthermore be regulated at the post-transcriptional level, e.g. by the presence of an antisense mRNA hybridizing to the rmPMT mRNA.
  • rmPMTs recombinant mutant PMT proteins derived from toxA by a single site-directed mutagenesis causing amino acid substitution, insertion or deletion, or by fusion of a subsequence of toxA to lacZ.
  • Whole-cell extracts containing different rmPMTs were assayed for their PE on NIH3T3 cells;
  • Fig. 5 shows a Coomassie Brilliant Blue stained SDS-PAGE gel showing protein profiles of 4HX-, derivative O-, and rPMT producing E. coli MC1000 (dam). Culture samples were taken in late exponential (lx), between late exponential and stationary (lxs), in early stationary (es), and late stationary (Is) growth phase. The relative amount of rmPMT or rPMT in the sample is indicated below each lane;
  • Fig. 11 shows SDS-PAGE analysis of fractions collected during Q-Sepharose chromatography of crude E. coli dO-containing extract. The 10% polyacrylamide gels were silver stained.
  • Fig. 25 shows the stability of derivative O enriched by Q- Sepharose and Phenyl Sepharose chromatography, during incubation with trypsin.
  • Fig. 32 shows SDS-PAGE analysis of purified ⁇ H1223. Two-fold dilutions of a preparation of purified ⁇ H1223 were analyzed by SDS-PAGE (10% gel, silver staining). Lanes 1 and 8: molecular weight markers; lanes 2-7: purified ⁇ H1223 diluted 2x, 4x, 8x, 16x, 32x and 64x, respectively.
  • the diluted extracts were used in the mitogenicity assay, which was carried out essentially as described by Rozengurt et al. (Rozengurt, E., T. Higgins, N. Chanter, A. J. Lax, and J. M. Staddon. 1990. Pasteurella multocida toxin: potent mitogen for cultured fibroblasts. Proc. Natl. Acad. Sci.

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  • Chemical & Material Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Organic Chemistry (AREA)
  • Medicinal Chemistry (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Biochemistry (AREA)
  • Biophysics (AREA)
  • General Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Molecular Biology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Communicable Diseases (AREA)
  • Peptides Or Proteins (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Preparation Of Compounds By Using Micro-Organisms (AREA)

Abstract

Cette invention concerne de nouvelles protéines mutantes recombinées qui sont capables de se lier à des anticorps dirigés contre la toxine Pasteurella multocida ainsi que leurs utilisations comme vaccins conférant une protection contre des maladies provoquées par Pasteurella multocida.
PCT/DK1993/000312 1992-09-30 1993-09-28 Proteine mutante recombinee derivee de pasteurella multocida et son procede de preparation WO1994008011A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
AU51081/93A AU5108193A (en) 1992-09-30 1993-09-28 Recombinant mutant pasteurella multocida protein and process for preparing the same

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
DK1207/92 1992-09-30
DK921207A DK120792D0 (da) 1992-09-30 1992-09-30 Protein
CN93114440.XA CN1091773A (zh) 1992-09-30 1993-09-30 重组突变出血败血性巴斯德氏菌蛋白及其制备方法

Publications (1)

Publication Number Publication Date
WO1994008011A1 true WO1994008011A1 (fr) 1994-04-14

Family

ID=36808690

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/DK1993/000312 WO1994008011A1 (fr) 1992-09-30 1993-09-28 Proteine mutante recombinee derivee de pasteurella multocida et son procede de preparation

Country Status (4)

Country Link
CN (1) CN1091773A (fr)
AU (1) AU5108193A (fr)
DK (1) DK120792D0 (fr)
WO (1) WO1994008011A1 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP1657248A3 (fr) * 2004-08-20 2006-05-24 National Chung-Hsing University Vaccin pour la prevention et le traitement de la rhinite atrophique progressive porcine

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1989009617A1 (fr) * 1988-04-12 1989-10-19 Nordisk Droge & Kemikalie A/S Vaccin de pasteurella
WO1991000100A2 (fr) * 1989-06-29 1991-01-10 Institute For Animal Health Limited Utilisations d'une toxine

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1989009617A1 (fr) * 1988-04-12 1989-10-19 Nordisk Droge & Kemikalie A/S Vaccin de pasteurella
WO1991000100A2 (fr) * 1989-06-29 1991-01-10 Institute For Animal Health Limited Utilisations d'une toxine

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
S.K. PETERSEN ET AL.: "Recombinant derivatives of Pasteurella multocida toxin: Candidates for a vaccine against progressive atrophic rhinitis", INFECTION AND IMMUNITY, vol. 59, no. 4, April 1991 (1991-04-01), WASHINGTON US, pages 1387 - 1393 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP1657248A3 (fr) * 2004-08-20 2006-05-24 National Chung-Hsing University Vaccin pour la prevention et le traitement de la rhinite atrophique progressive porcine

Also Published As

Publication number Publication date
CN1091773A (zh) 1994-09-07
DK120792D0 (da) 1992-09-30
AU5108193A (en) 1994-04-26

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