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Moraxella (branhamella) catarrhalis polypeptides and corresponding DNA fragments

A DNA sequence and fragment technology, applied in the field of SMC-1 and SMC-2 polypeptides of Moraxella catarrhalis

Inactive Publication Date: 2004-11-24
ID BIOMEDICAL
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This protein is highly conserved among strains of Moraxella catarrhalis, yet adults with chronic obstructive pulmonary disease show differences in the immune response to OMP CD
[0005] There is therefore still an unmet need for M. catarrhalis polypeptides for the prevention, diagnosis and / or treatment of M. catarrhalis (Brahamella) infection

Method used

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  • Moraxella (branhamella) catarrhalis polypeptides and corresponding DNA fragments
  • Moraxella (branhamella) catarrhalis polypeptides and corresponding DNA fragments

Examples

Experimental program
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Embodiment 1

[0176] This example illustrates the cloning and molecular characterization of the SMC-1 gene and the corresponding polypeptide.

[0177] Moraxella catarrhalis SMC-1 (SEQ ID NO: 1) was amplified from Moraxella catarrhalis strain ETSU C-2 genomic DNA by PCR method (DNAThermal Cycler GeneAmp PCR system 2400 Perkin Elmer, San Jose, CA) The coding region was amplified using oligonucleotides containing base extensions for the addition of restriction sites NcoI (CCATGG) and XhoI (CTCGAG): RIOS30 (5'-TATGTACCATGGCTGAACTCAATACCAGCCGTTCA-3') and RIOS31 ( 5'-GGCATGCTCGAGGTAATCATGTCTCCAAGCATTTTG-3'). PCR products were purified from agarose gels using the QIAquick Gel Extraction Kit (Qiagen, Chatsworth, CA) according to the manufacturer's instructions and digested with NcoI and XhoI (Amersham PharmaciaBiotech, Inc, Baie d'Urfé, Canada). The pET21d(+) vector (Novagen, Madison, WI) was digested with Ncol and XhoI and the product was purified from agarose gel using the QIAquick Gel Extractio...

Embodiment 2

[0183] This example illustrates the cloning and molecular characterization of the SMC-2 gene and the corresponding polypeptide. Moraxella catarrhalis SMC-2 (SEQ ID NO: 3) was amplified by PCR method (DNA ThermalCycler GeneAmp PCR system 2400 Perkin Elmer, San Jose, CA) from Moraxella catarrhalis strain ETSU C-2 genomic DNA For the coding region, amplified using oligonucleotides containing a base extension for the addition of restriction sites NcoI (CCATGG) and XhoI (CTCGAG): RIOS20 and RIOS21, both of which are listed in Table 1 out. The method for cloning the SMC-2 gene into an expression vector and performing sequencing is similar to the method described in Example 1.

[0184] It has been determined that the open reading frame (ORF) encoding SMC-2 contains 957-bp (base pairs) and encodes a polypeptide with 318 amino acid residues. The predicted pI is 5.78 and the predicted molecular weight is 35954.10Da. Using Spscan software (Wisconsin Sequence AnalysisiPackage; Genetics ...

Embodiment 3

[0187] This example illustrates the cloning of M. catarrhalis genes in the CMV plasmid pCMV-GH.

[0188] In-phase insertion of the DNA coding region of the Moraxella catarrhalis polypeptide into the vector pCMV-GH (Tang et al., Nature, 1992, 356:152) Human growth under the transcriptional control of the plasmid cytomegalovirus (CMV) promoter Downstream of the hormone (hGH) gene. The CMV promoter was not functional in E. coli cells, but was active when administered to eukaryotic cells. The ampicillin resistance gene was also inserted into the vector.

[0189] From the genomic DNA of Moraxella catarrhalis strain ETSU C-2, the SMC-1 (SEQ ID NO: 1) and SMC-2 (SEQ ID NO: 3) coding regions without the leader peptide were amplified by PCR to amplify Oligonucleotides containing a base extension for the addition of restriction sites BamHI (GGATCC), BglII (AGATCT), SalI (GTCGAC) or HindIII (AAGCTT) were used, listed in Table 1. PCR products were purified from agarose gels using the Q...

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Abstract

The present invention relates to polypeptides of Moraxella (Branhamela) catarrhalis which may be used for prophy-laxis, diagnostic and / or therapy purposes.

Description

field of invention [0001] The present invention relates to a polypeptide that can be used for preventing, diagnosing and / or treating Moraxella (Branhamella) catarrhalis infection, especially the polypeptide of Moraxella (Branhamella) catarrhalis SMC-1 and SMC-2 polypeptides. Background of the invention [0002] Moraxella catarrhalis (Brahamella) is a Gram-negative diplococcus that can cause respiratory infections in humans. Moraxella catarrhalis is currently considered to be the third most common cause of otitis media in infants and children, after Streptococcus pneumoniae and Haemophilus influenzae. M. catarrhalis has also been associated with several other types of infection, including sinusitis, persistent cough, acute pharyngitis, suppurative keratitis, and neonatal conjunctivitis. [0003] Since nearly 90% of M. catarrhalis strains are antibiotic resistant (β-lactamase positive) and have a high incidence of recurrent otitis media, it is necessary to develop a strain t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/53A61K38/00A61K39/00A61P11/02A61P11/04A61P27/02A61P27/16A61P31/00A61P37/02C07K14/195C07K14/21C07K19/00C12N1/15C12N1/19C12N1/21C12N5/10C12N15/09C12N15/31C12P21/02G01N33/569
CPCY10S435/975C07K14/212A61K39/00A61P11/02A61P11/04A61P11/14A61P27/02A61P27/16A61P31/00A61P37/02A61P37/04
Inventor D·马丁J·哈梅尔B·R·布罗德尔S·理欧克斯J·库图尔
Owner ID BIOMEDICAL
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