Acanthamoeba alternative oxidase genes: identification, characterisation and potential as antimicrobial targets

Fiona L. Henriquez, James McBride, Sara J. Campbell, Tania Ramos, Paul R. Ingram, F. Roberts, Sinead Tinney, C.W. Roberts

Research output: Contribution to journalArticle

9 Citations (Scopus)

Abstract

Alternative oxidase (AOX) is a mitochondrial protein that acts as an alternative terminal oxidase to the conventional cytochrome oxidases. It is present in certain prokaryotes, plants, fungi and some protozoa but absent in mammals. AOX activity has previously been described in Acanthamoeba, although no genetic evidence has been reported. Herein, two AOX (AcAOX) genes designated isoforms A and B, were obtained from Acanthamoeba castellanii by a combination of degenerate PCR from cDNA and a series of 5′ and 3′ rapid amplification of cDNA ends. The corresponding genomic sequences of these AcAOXs were also obtained. Each gene spans six exons over a region of 1607 and 1619 bp, respectively. Isoforms A and B have open reading frames of 1113 and 1125 bp, respectively. Each encodes a protein with a predicted molecular weight of 42 kDa. Each AcAOX protein has a predicted cleavable mitochondrial targeting sequence. The full-length AcAOX is functionally active as it complements hemL-deficient Escherichia coli and inhibited by the inhibitor of AOX, salicylhydroxamic acid (SHAM). SHAM is effective against A. castellanii and Acanthamoeba polyphaga only when used in conjunction with antimycin A, an inhibitor of the conventional cytochrome respiratory pathway. Transcripts for AcAOX are increased during the encystment process, indicating a possible role for alternative respiration during stress.
LanguageEnglish
Pages1417-1424
Number of pages8
JournalInternational Journal for Parasitology
Volume39
Issue number13
DOIs
Publication statusPublished - Nov 2009

Fingerprint

Acanthamoeba
Acanthamoeba castellanii
Genes
Protein Isoforms
Complementary DNA
Antimycin A
Mitochondrial Proteins
Electron Transport Complex IV
Cytochromes
Open Reading Frames
Mammals
Exons
Respiration
Proteins
Fungi
Molecular Weight
Escherichia coli
Polymerase Chain Reaction
alternative oxidase
salicylhydroxamic acid

Keywords

  • acanthamoeba
  • animals
  • anti-infective agents
  • antiprotozoal agents
  • drug discovery
  • humans
  • mitochondria
  • mitochondrial proteins
  • molecular sequence data
  • oxidoreductases
  • plant proteins
  • polymerase chain reaction
  • salicylamides

Cite this

Henriquez, Fiona L. ; McBride, James ; Campbell, Sara J. ; Ramos, Tania ; Ingram, Paul R. ; Roberts, F. ; Tinney, Sinead ; Roberts, C.W. / Acanthamoeba alternative oxidase genes : identification, characterisation and potential as antimicrobial targets. In: International Journal for Parasitology. 2009 ; Vol. 39, No. 13. pp. 1417-1424.
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Acanthamoeba alternative oxidase genes : identification, characterisation and potential as antimicrobial targets. / Henriquez, Fiona L.; McBride, James; Campbell, Sara J.; Ramos, Tania; Ingram, Paul R.; Roberts, F.; Tinney, Sinead; Roberts, C.W.

In: International Journal for Parasitology, Vol. 39, No. 13, 11.2009, p. 1417-1424.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Acanthamoeba alternative oxidase genes

T2 - International Journal for Parasitology

AU - Henriquez, Fiona L.

AU - McBride, James

AU - Campbell, Sara J.

AU - Ramos, Tania

AU - Ingram, Paul R.

AU - Roberts, F.

AU - Tinney, Sinead

AU - Roberts, C.W.

PY - 2009/11

Y1 - 2009/11

N2 - Alternative oxidase (AOX) is a mitochondrial protein that acts as an alternative terminal oxidase to the conventional cytochrome oxidases. It is present in certain prokaryotes, plants, fungi and some protozoa but absent in mammals. AOX activity has previously been described in Acanthamoeba, although no genetic evidence has been reported. Herein, two AOX (AcAOX) genes designated isoforms A and B, were obtained from Acanthamoeba castellanii by a combination of degenerate PCR from cDNA and a series of 5′ and 3′ rapid amplification of cDNA ends. The corresponding genomic sequences of these AcAOXs were also obtained. Each gene spans six exons over a region of 1607 and 1619 bp, respectively. Isoforms A and B have open reading frames of 1113 and 1125 bp, respectively. Each encodes a protein with a predicted molecular weight of 42 kDa. Each AcAOX protein has a predicted cleavable mitochondrial targeting sequence. The full-length AcAOX is functionally active as it complements hemL-deficient Escherichia coli and inhibited by the inhibitor of AOX, salicylhydroxamic acid (SHAM). SHAM is effective against A. castellanii and Acanthamoeba polyphaga only when used in conjunction with antimycin A, an inhibitor of the conventional cytochrome respiratory pathway. Transcripts for AcAOX are increased during the encystment process, indicating a possible role for alternative respiration during stress.

AB - Alternative oxidase (AOX) is a mitochondrial protein that acts as an alternative terminal oxidase to the conventional cytochrome oxidases. It is present in certain prokaryotes, plants, fungi and some protozoa but absent in mammals. AOX activity has previously been described in Acanthamoeba, although no genetic evidence has been reported. Herein, two AOX (AcAOX) genes designated isoforms A and B, were obtained from Acanthamoeba castellanii by a combination of degenerate PCR from cDNA and a series of 5′ and 3′ rapid amplification of cDNA ends. The corresponding genomic sequences of these AcAOXs were also obtained. Each gene spans six exons over a region of 1607 and 1619 bp, respectively. Isoforms A and B have open reading frames of 1113 and 1125 bp, respectively. Each encodes a protein with a predicted molecular weight of 42 kDa. Each AcAOX protein has a predicted cleavable mitochondrial targeting sequence. The full-length AcAOX is functionally active as it complements hemL-deficient Escherichia coli and inhibited by the inhibitor of AOX, salicylhydroxamic acid (SHAM). SHAM is effective against A. castellanii and Acanthamoeba polyphaga only when used in conjunction with antimycin A, an inhibitor of the conventional cytochrome respiratory pathway. Transcripts for AcAOX are increased during the encystment process, indicating a possible role for alternative respiration during stress.

KW - acanthamoeba

KW - animals

KW - anti-infective agents

KW - antiprotozoal agents

KW - drug discovery

KW - humans

KW - mitochondria

KW - mitochondrial proteins

KW - molecular sequence data

KW - oxidoreductases

KW - plant proteins

KW - polymerase chain reaction

KW - salicylamides

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DO - 10.1016/j.ijpara.2009.04.011

M3 - Article

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SP - 1417

EP - 1424

JO - International Journal for Parasitology

JF - International Journal for Parasitology

SN - 0020-7519

IS - 13

ER -