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Molecular Cloning and Characterization of a Gene Encoding a 13.1 kDa Antigenic Protein of Naegleria fowleri
Author(s) -
SHIN HOJOON,
CHO MYUNGSOO,
JUNG SUKYUL,
KIM HYUNGIL,
PARK SUN,
KIM HANJIP,
IM KYUNGIL
Publication year - 2001
Publication title -
journal of eukaryotic microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.067
H-Index - 77
eISSN - 1550-7408
pISSN - 1066-5234
DOI - 10.1111/j.1550-7408.2001.tb00211.x
Subject(s) - biology , immunoscreening , naegleria fowleri , microbiology and biotechnology , peptide sequence , gene , naegleria , nucleic acid sequence , complementary dna , molecular cloning , recombinant dna , sequence analysis , amino acid , biochemistry , cdna library , genetics , protozoa , virology , meningoencephalitis
. An antigen‐related gene was cloned from a cDNA expression library of Naegleria fowleri by immunoscreening with sera obtained from mice that were either immunized with an amoebic lysate or infected with trophozoites. The coding nucleotide sequence of the cloned gene consisted of 357 bases that were translated into 119 amino acids. This gene was designated as nfal. The predicted amino acid sequence of Nfal protein has two potential glycosylation and three potential phosphorylation sites, and its predicted secondary structure consists of four helices and three corners. The deduced amino acid sequence of Nfal protein shares 43% identity with the myohemerythrin (myoHr) protein from a marine annelid, Nereis diversicolor , including 100% identity in conserved regions and iron‐binding residues. A phylogenetic tree constructed from amino acid sequences placed the N. fowleri Nfal protein outside of a cluster of myoHr proteins from eight invertebrates. A purified recombinant protein that migrated as a 13.1 kDa species in SDS‐PAGE was produced. This recombinant protein exhibited a strong immunoreactivity with infected, immune, and anti‐Nfal sera. In addition. an anti‐Nfal serum reacted with an amoeba lysate in immunoblotting analysis. The present nfal gene encoding the myoHr‐like protein is the first myoHr gene cloned from protozoa, and the Nfal antigen may be useful in diagnostic studies.