Targeting and subcellular localization of Toxoplasma gondii catalase. Identification of peroxisomes in an apicomplexan parasite

Achim J. Kaasch, Keith A Joiner

Research output: Contribution to journalArticle

36 Citations (Scopus)

Abstract

We sought to identify and characterize peroxisomes in the apicomplexan parasite Toxoplasma gondii. To initiate this process, we first cloned and sequenced the gene for T. gondii catalase (EC 1.11.1.6), a marker enzyme for peroxisomes in eukaryotic cells. The gene predicts a protein of 57.2 kDa and 502 amino acids and has a strong homology to other eukaryotic catalases. A polyclonal antiserum raised against a glutathione S-transferase fusion protein recognized a single band with a molecular mass of 63 kDa by immunoblot. By immunofluorescence T. gondii catalase is present primarily in a punctate staining pattern anterior to the parasite nucleus. This compartment is distinguishable from other parasite organelles, namely micronemes, rhoptries, dense granules, and the apicoplast. Cytochemical visualization of catalase using diaminobenzidine precipitation gives a vesicular staining pattern anterior to the nucleus at the light level and round, vesicular structures with an estimated diameter of 100-300 nm by electron microscopy. T. gondii catalase has a putative C-terminal peroxisomal targeting signal in the last 3 amino acids (-AKM). Expression of T. gondii catalase in mammalian cells results in peroxisomal localization, whereas a construct lacking the targeting signal remains in the cytosol. Furthermore, addition of -AKM to the C terminus of chloramphenicol acetyltransferase is sufficient to target this protein to peroxisomes. These results provide the first evidence for peroxisomes in Apicomplexan parasites.

Original languageEnglish (US)
Pages (from-to)1112-1118
Number of pages7
JournalJournal of Biological Chemistry
Volume275
Issue number2
DOIs
StatePublished - Jan 14 2000
Externally publishedYes

Fingerprint

Peroxisomes
Toxoplasma
Catalase
Parasites
Apicoplasts
Genes
Staining and Labeling
Amino Acids
Proteins
Chloramphenicol O-Acetyltransferase
Molecular mass
Eukaryotic Cells
Glutathione Transferase
Organelles
Cytosol
Electron microscopy
Fluorescent Antibody Technique
Immune Sera
Electron Microscopy
Fusion reactions

ASJC Scopus subject areas

  • Biochemistry

Cite this

Targeting and subcellular localization of Toxoplasma gondii catalase. Identification of peroxisomes in an apicomplexan parasite. / Kaasch, Achim J.; Joiner, Keith A.

In: Journal of Biological Chemistry, Vol. 275, No. 2, 14.01.2000, p. 1112-1118.

Research output: Contribution to journalArticle

@article{b52b9a8189f44a87ba3aa6e6fd220809,
title = "Targeting and subcellular localization of Toxoplasma gondii catalase. Identification of peroxisomes in an apicomplexan parasite",
abstract = "We sought to identify and characterize peroxisomes in the apicomplexan parasite Toxoplasma gondii. To initiate this process, we first cloned and sequenced the gene for T. gondii catalase (EC 1.11.1.6), a marker enzyme for peroxisomes in eukaryotic cells. The gene predicts a protein of 57.2 kDa and 502 amino acids and has a strong homology to other eukaryotic catalases. A polyclonal antiserum raised against a glutathione S-transferase fusion protein recognized a single band with a molecular mass of 63 kDa by immunoblot. By immunofluorescence T. gondii catalase is present primarily in a punctate staining pattern anterior to the parasite nucleus. This compartment is distinguishable from other parasite organelles, namely micronemes, rhoptries, dense granules, and the apicoplast. Cytochemical visualization of catalase using diaminobenzidine precipitation gives a vesicular staining pattern anterior to the nucleus at the light level and round, vesicular structures with an estimated diameter of 100-300 nm by electron microscopy. T. gondii catalase has a putative C-terminal peroxisomal targeting signal in the last 3 amino acids (-AKM). Expression of T. gondii catalase in mammalian cells results in peroxisomal localization, whereas a construct lacking the targeting signal remains in the cytosol. Furthermore, addition of -AKM to the C terminus of chloramphenicol acetyltransferase is sufficient to target this protein to peroxisomes. These results provide the first evidence for peroxisomes in Apicomplexan parasites.",
author = "Kaasch, {Achim J.} and Joiner, {Keith A}",
year = "2000",
month = "1",
day = "14",
doi = "10.1074/jbc.275.2.1112",
language = "English (US)",
volume = "275",
pages = "1112--1118",
journal = "Journal of Biological Chemistry",
issn = "0021-9258",
publisher = "American Society for Biochemistry and Molecular Biology Inc.",
number = "2",

}

TY - JOUR

T1 - Targeting and subcellular localization of Toxoplasma gondii catalase. Identification of peroxisomes in an apicomplexan parasite

AU - Kaasch, Achim J.

AU - Joiner, Keith A

PY - 2000/1/14

Y1 - 2000/1/14

N2 - We sought to identify and characterize peroxisomes in the apicomplexan parasite Toxoplasma gondii. To initiate this process, we first cloned and sequenced the gene for T. gondii catalase (EC 1.11.1.6), a marker enzyme for peroxisomes in eukaryotic cells. The gene predicts a protein of 57.2 kDa and 502 amino acids and has a strong homology to other eukaryotic catalases. A polyclonal antiserum raised against a glutathione S-transferase fusion protein recognized a single band with a molecular mass of 63 kDa by immunoblot. By immunofluorescence T. gondii catalase is present primarily in a punctate staining pattern anterior to the parasite nucleus. This compartment is distinguishable from other parasite organelles, namely micronemes, rhoptries, dense granules, and the apicoplast. Cytochemical visualization of catalase using diaminobenzidine precipitation gives a vesicular staining pattern anterior to the nucleus at the light level and round, vesicular structures with an estimated diameter of 100-300 nm by electron microscopy. T. gondii catalase has a putative C-terminal peroxisomal targeting signal in the last 3 amino acids (-AKM). Expression of T. gondii catalase in mammalian cells results in peroxisomal localization, whereas a construct lacking the targeting signal remains in the cytosol. Furthermore, addition of -AKM to the C terminus of chloramphenicol acetyltransferase is sufficient to target this protein to peroxisomes. These results provide the first evidence for peroxisomes in Apicomplexan parasites.

AB - We sought to identify and characterize peroxisomes in the apicomplexan parasite Toxoplasma gondii. To initiate this process, we first cloned and sequenced the gene for T. gondii catalase (EC 1.11.1.6), a marker enzyme for peroxisomes in eukaryotic cells. The gene predicts a protein of 57.2 kDa and 502 amino acids and has a strong homology to other eukaryotic catalases. A polyclonal antiserum raised against a glutathione S-transferase fusion protein recognized a single band with a molecular mass of 63 kDa by immunoblot. By immunofluorescence T. gondii catalase is present primarily in a punctate staining pattern anterior to the parasite nucleus. This compartment is distinguishable from other parasite organelles, namely micronemes, rhoptries, dense granules, and the apicoplast. Cytochemical visualization of catalase using diaminobenzidine precipitation gives a vesicular staining pattern anterior to the nucleus at the light level and round, vesicular structures with an estimated diameter of 100-300 nm by electron microscopy. T. gondii catalase has a putative C-terminal peroxisomal targeting signal in the last 3 amino acids (-AKM). Expression of T. gondii catalase in mammalian cells results in peroxisomal localization, whereas a construct lacking the targeting signal remains in the cytosol. Furthermore, addition of -AKM to the C terminus of chloramphenicol acetyltransferase is sufficient to target this protein to peroxisomes. These results provide the first evidence for peroxisomes in Apicomplexan parasites.

UR - http://www.scopus.com/inward/record.url?scp=0033983257&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0033983257&partnerID=8YFLogxK

U2 - 10.1074/jbc.275.2.1112

DO - 10.1074/jbc.275.2.1112

M3 - Article

C2 - 10625653

AN - SCOPUS:0033983257

VL - 275

SP - 1112

EP - 1118

JO - Journal of Biological Chemistry

JF - Journal of Biological Chemistry

SN - 0021-9258

IS - 2

ER -