Molecular cloning and characterization of a cytosolic heat shock protein 70 from Naegleria fowleri.
A gene encoding a cytosolic heat shock protein 70 from pathogenic Naegleria fowleri (Nf-cHSP70) was identified. The Nf-cHSP70 was 2,062 bp in length with an open reading frame of 1,980 bp encoding 659 amino acid residues. The deduced amino acid sequence of the gene shared high sequence identities with HSP70s from other parasitic organisms and mammals. The characteristic domains, including N-terminal ATPase domain, calmodulin-binding domain, and EE(D)VD motif, found in HSP70s were also well conserved in this gene. The recombinant Nf-cHSP70 protein showed strong antigenicity against the sera from mice experimentally infected with N. fowleri. Immunofluorescence assay showed that Nf-cHSP70 localized in cytosol of the parasite. The results from semi-quantitative RT-PCR and Western blot analyses demonstrated the expression levels of gene transcripts, and its products were significantly increased at high temperature (42 degrees C). The definitive biological roles of Nf-cHSP70 are not clear, but it may protect the parasite under environmental changes especially high temperature.
Song
Kyoung-Ju
KJ
Department of Microbiology, Ajou University School of Medicine, Suwon 443-721, South Korea.
Song
Kyung-Hui
KH
Na
Byoung-Kuk
BK
Kim
Jong-Hyun
JH
Kwon
Daeho
D
Park
Sun
S
Pak
Jhang-Ho
JH
Im
Kyung-Il
KI
Shin
Ho-Joon
HJ
D020816:Amino Acid Motifs / Q000235:genetics; D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D017124:Conserved Sequence; D003600:Cytosol / Q000737:chemistry; D016054:DNA, Protozoan / Q000737:chemistry / Q000235:genetics; D005786:Gene Expression Regulation; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics*; D008856:Microscopy, Fluorescence; D008969:Molecular Sequence Data; D016848:Naegleria fowleri / Q000235:genetics*; D016366:Open Reading Frames; D017434:Protein Structure, Tertiary / Q000235:genetics; D015800:Protozoan Proteins / Q000235:genetics*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D016053:RNA, Protozoan / Q000096:biosynthesis / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D013696:Temperature
D016054:DNA, Protozoan; D018840:HSP70 Heat-Shock Proteins; D015800:Protozoan Proteins; D012333:RNA, Messenger; D016053:RNA, Protozoan