Journal of Molecular Biology
Volume 325, Issue 2, 10 January 2003, Pages 259-274
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Characterization of the ABCA Transporter Subfamily: Identification of Prokaryotic and Eukaryotic Members, Phylogeny and Topology

https://doi.org/10.1016/S0022-2836(02)01105-1Get rights and content

Abstract

An alignment of the mammalian ABCA transporters enabled the identification of sequence segments, specific to the ABCA subfamily, which were used as queries to search for eukaryotic and prokaryotic homologues. Thirty-seven eukaryotic half and full-length transporters were found, and a close relationship with prokaryotic subfamily 7 transporters was detected. Each half of the ABCA full-transporters is predicted to comprise a membrane-spanning domain (MSD) composed of six helices and a large extracellular loop, followed by a nucleotide-binding domain (NBD) and a conserved cytoplasmic 80-residue sequence, which might have a regulatory function. The topology predicted for the ABCA transporters was compared to the crystal structures of the MsbA and BtuCD bacterial transporters. The alignment of the MSD and NBD domains provided an estimate of the degree of residue conservation in the cytoplasmic, extracellular and transmembrane domains of the ABCA transporter subfamily. The phylogenic tree of eukaryotic ABCA transporters based upon the NBD sequences, consists of three major clades, corresponding to the half-transporter single NBDs and to the full-transporter NBDls and NBD2s. A phylogenic tree of prokaryotic transporters and the eukaryotic ABCA transporters confirmed the evolutionary relationship between prokaryotic subfamily 7 transporters and eukaryotic ABCA half and full-transporters.

Introduction

ATP-binding cassette (ABC) transporters form a large protein family, and are well represented in all species from prokaryotes to man. ABC transporters bind and hydrolyse ATP, and utilise the energy of ATP hydrolysis to mediate the translocation of a wide variety of substances across cellular membranes. Forty-eight human ABC transporters have been identified, and subdivided into seven phylogenic groupings, subfamily ABCA through to ABCG†.1., 2. Several ABC transporters are linked to genetic diseases such as ABCA1 to Tangier disease and HDL deficiency, ABCA4 to Stargardt disease, ABCC2 to Dubin–Johnson syndrome, ABCC7 (CFTR) to cystic fibrosis, ABCD1 to adrenoleukodystrophy, ABCG5 and ABCG8 to sitosterolemia.3

A functional ABC transporter comprises two membrane spanning domains (MSD1 and MSD2), which form a single transmembrane pore through which the substrate is presumably translocated, and two cytoplasmic nucleotide-binding domains (NBDl and NBD2).2 MSD domains are predicted to contain six to ten transmembrane helices, consistent with a high resolution structure of MsbA and BtuC from Escherichia coli.4., 5. The NBDs contain the Walker A/B motif together with the ABC signature motif.1 Certain ABC transporters, referred to as full-transporters, consist of a single polypeptide, while others are formed by two half-transporters, as is the case for MsbA of E. coli, which has high sequence similarity with the mammalian P-glycoproteins of subfamily B. Half-transporters contain one MSD and one NBD, and assemble as functional homo- or heterodimers.6

The human ABCA transporter subfamily consists of 12 members.7 Among these, the putative lipid transporter ABCA1 plays a key metabolic role in reverse cholesterol transport from peripheral tissues to the liver, and in HDL synthesis.6 Mutations in human ABCA1 were described in patients with Tangier disease,8 with familial hypoalphalipoproteinemia, and early-onset development of atherosclerosis. ABCA4 is a putative transporter of retinoids in rod cells, and mutations in the ABCA4 gene cause retinal degenerative disorders such as Stargardt disease, age-related macular dystrophy, retinitis pigmentosa RP19, and cone rod dystrophy.9 ABCA2 is a lysosome-associated transporter found in brain oligodendrocytes.10 Human ABCA3 is expressed in the lung alveolar type II cells, while ABCA7 is expressed mainly in the spleen and in hematopoietic tissues.6 A cluster of five new members of the ABCA family was recently identified on chromosome 17.7., 11. These transporters, named ABCA5, 6, 8, 9 and 10 are expressed in skeletal muscle (ABCA5 and 10), liver (ABCA6), heart (ABCA9), and ovary (ABCA8).7 The functional properties and substrate specificity of the ABCA12 transporter are still unknown.7 Putative homologues of the human ABCA subfamily members were identified in Drosophila melanogaster, Caenorhabditis elegans and in Arabidopsis thaliana.3., 12. The ced7 transporter in C. elegans was proposed as the orthologue of human ABCA1 since both transporters seem involved in the phagocytic engulfment of apoptotic cell bodies and translocation of phosphatidylserine.13

Here, we identified new members of the ABCA subfamily in different eukaryotic species, and ABCA homologues in prokaryotes. Multiple sequence alignments of ABCA transporters enabled the study of the phylogeny and evolution of the ABCA subfamily. These multiple alignments provide better insight into the conservation, and domain organization of the ABCA transporters, and will facilitate future studies of the ABCA structure–function relationship.

Section snippets

Internal symmetry within the ABCA full transporters

Here, we first identified putative new members of the ABCA subfamily among the eukaryotic and prokaryotic species and confirmed that they belong to this ABC subfamily. The search for new ABCA homologues first required the definition of query sequences that are specific to the ABCA subfamily only. These query sequences were then used in a combination of the SSEARCH, PSI-BLAST and SAM-t99 search algorithms, in order to detect a maximal number of ABCA homologues.14 In order to detect both

Discussion

The sequence alignment of 17 mammalian ABCA transporters, enabled the identification of sequence segments specific to the ABCA subfamily, which were used to search for new ABCA transporters. Using PSI-BLAST, SSEARCH and SAM-t99 Hidden Markov Models, we identified 37 eukaryotic ABCA transporters, including both half and full-ABCA transporters, demonstrated the high internal symmetry in ABCA full-transporters and proposed a general topology for the ABCA subfamily. We also detected prokaryotic

Materials and Methods

Mammalian ABCA transporter sequences were aligned with DIALIGN,33 and CLUSTALX,15 using the BLOSUM substitution matrix series with a gap opening penalty of 5, and a gap extension penalty of 0.05. Residue conservation calculations along the sequences helped identify conserved segments, which were used as query sequences, Q1, Q2, Q3, to search for mammalian ABCA homologues. These were detected using SSEARCH,34 PSI-BLAST (–B option),35 combined with the OCTOPUS editing tool, and SAM-t9936

Acknowledgements

This work was supported by Grant of the Funds for Scientific Research-Flanders 3G032202. S.R. is a recipient of a doctoral Research Grant of the University Gent (011V0201-VEO). We are grateful to Dr C. Shoulders (London) for her comments and suggestions about the manuscript.

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