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Homology building as a means to define antigenic epitopes on dihydrofolate reductase (DHFR) from Plasmodium falciparum

DOI: 10.1186/1475-2875-3-16

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Abstract:

A homology model of Pf-DHFR domain was employed to define an epitope for the development of site-specific antibodies against Pf-DHFR. The homology model suggested an exposed loop encompassing amino acid residues 64–100. A synthetic peptide of 37-mers whose sequence corresponded to the sequence of amino acid residues 64–100 of Pf-DHFR was synthesized and used to immunize mice for antibodies. Additionally, polyclonal antibodies recognizing a recombinant DHFR enzyme were produced in rabbits.Serum from mice immunized with the 37-mer showed strong reactivity against both the immunizing peptide, recombinant DHFR and a preparation of crude antigen from P. falciparum infected red blood cells. Five monoclonal antibodies were obtained, one of which showed reactivity towards crude antigen prepared from P. falciparum infected red cells. Western blot analysis revealed that both the polyclonal and monoclonal antibodies recognized Pf-DHFR. Our study provides insight into the potential use of homology models in general and of Pf-DHFR in particular in predicting antigenic malarial surface epitopes.Antibodies raised against short peptide fragments of a given protein have been reported to be able to cross-react with the native protein [1]. The identification of peptide epitopes simulating the native protein has traditionally been based on amino acid sequences or sequence motifs exposed on the outer surface of the protein structure, thereby making these peptides potential candidates as antigen epitopes. Examples of algorithms for selecting and defining properties of exposed peptide sequences include plots of hydrophilicity, hydrophobicity, external flexibility and antigenic index. However, these algorithms provide only crude approximations of the native structures, and antibodies raised against the selected peptides are often lacking reactivity or show low degree of cross-reactivity with the native protein [2]. In recent years, the number of proteins for which three-dimensional structu

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