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ACBP4 and ACBP5, novel Arabidopsis acyl-CoA-binding proteins with kelch motifs that bind oleoyl-CoA

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Abstract

In plants, fatty acids synthesized in the chloroplasts are exported as acyl-CoA esters to the endoplasmic reticulum (ER). Cytosolic 10-kDa acyl-CoA-binding proteins (ACBPs), prevalent in eukaryotes, are involved in the storage and intracellular transport of acyl-CoAs. We have previously characterized Arabidopsis thaliana cDNAs encoding membrane-associated ACBPs with ankyrin repeats, designated ACBP1 and ACBP2, which show conservation to cytosolic ACBPs at the acyl-CoA-binding domain. Analysis of the Arabidopsis genome has revealed the presence of three more genes encoding putative proteins with acyl-CoA-binding domains, designated ACBP3, ACBP4 and ACBP5. Homologues of ACBP1 to ACBP5 have not been reported in any other organism. We show by reverse-transcriptase polymerase chain reaction (RTPCR) analysis that ACBP3, ACBP4 and ACBP5 are expressed in all plant organs, like ACBP1 and ACBP2. ACBP4 and ACBP5 that share 81.4% identity and which contain kelch motifs were further investigated. To demonstrate their function in binding acyl-CoA, we have expressed them as (His)6-tagged recombinant proteins in Escherichia coli for in vitro binding assays. Both (His)6-ACBP4 and (His)6-ACBP5 bind [14C]oleoyl-CoA with high affinity, [14C]palmitoyl-CoA with lower affinity and did not bind [14C]arachidonyl-CoA. Eight mutant forms of each protein with single amino acid substitutions within the acyl-CoA-binding domain were produced and analyzed. On binding assays, all mutants were impaired in oleoyl-CoA-binding. Hence, these novel ACBPs with kelch motifs have functional acyl-CoA-binding domains that bind oleoyl-CoA. Their predicted cytosol localization suggests that they could maintain an oleoyl-CoA pool in the cytosol or transport oleoyl-CoA from the plastids to the ER in plant lipid metabolism.

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  1. Hong-Ye Li

    Present address: South China Institute of Botany, Chinese Academy of Sciences, 510650, Guangzhou, China

Authors and Affiliations

  1. Department of Botany, The University of Hong Kong, Pokfulam Road, Hong Kong, China

    Ka-Chun Leung, Hong-Ye Li, Girish Mishra & Mee-Len Chye

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  1. Ka-Chun Leung
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Correspondence to Mee-Len Chye.

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Leung, KC., Li, HY., Mishra, G. et al. ACBP4 and ACBP5, novel Arabidopsis acyl-CoA-binding proteins with kelch motifs that bind oleoyl-CoA. Plant Mol Biol 55, 297–309 (2005). https://doi.org/10.1007/s11103-005-0642-7

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