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Cloning and sequencing of Coxiella burnetii outer membrane protein gene com1
L R Hendrix1, L P Mallavia, J E Samuel
1MicroCarb Inc., Gaithersburg, Maryland 20879.
Abstract:
The gene for an approximately 27-kDa outer membrane-associated, immunoreactive protein was cloned from the rickettsial pathogen Coxiella burnetii. The gene, designated com1 for Coxiella outer membrane protein 1, was expressed in Escherichia coli, presumably by its own promoter. The complete nucleotide sequence of the gene was determined. The deduced amino acid sequence of 252 residues includes a putative leader sequence. The leader sequence is recognized in and removed by E. coli on the basis of the difference in the molecular mass of the protein produced in an in vitro transcription-translation system (27.6 kDa) and that of the protein immunoprecipitated from an iodinated E. coli clone (25.7 kDa). The Com1 protein expressed in E. coli was proteinase K sensitive in nondisrupted cells and soluble in 1% Sarkosyl, suggesting a loose association with the outer membrane. While the complete predicted sequence of the Com1 protein does not show any overall similarity to those of previously described proteins, a region which includes the only two cysteines in Com1 is homologous to the catalytic site of protein disulfide oxidoreductases.
Insights
The Coxiella burnetii com1 gene was cloned and sequenced. The resulting outer membrane protein (Com1) shows homology to oxidoreductases, suggesting a potential role in rickettsial pathogenesis.
Area of Science:
- Microbiology
- Molecular Biology
- Immunology
Background:
- Coxiella burnetii is a rickettsial pathogen responsible for Q fever.
- Outer membrane proteins are crucial for bacterial interactions and virulence.
Purpose of the Study:
- To clone and characterize the gene encoding a major outer membrane protein of Coxiella burnetii.
- To investigate the properties and potential function of the Com1 protein.
Main Methods:
- Gene cloning and nucleotide sequencing of the com1 gene from Coxiella burnetii.
- Expression of the com1 gene in Escherichia coli.
- Analysis of protein expression, molecular mass, and localization.
Main Results:
- The com1 gene was successfully cloned and sequenced, yielding the Coxiella outer membrane protein 1 (Com1).
- Com1 protein expressed in E. coli exhibited characteristics of a loosely outer membrane-associated protein.
- Sequence analysis revealed a region homologous to the catalytic site of protein disulfide oxidoreductases.
Conclusions:
- The Com1 protein is a novel outer membrane protein of Coxiella burnetii with potential enzymatic activity.
- Understanding Com1's function may provide insights into Coxiella burnetii pathogenesis and host interactions.