Diversity and dispersal of a ubiquitous protein family: acyl-CoA dehydrogenases

Abstract

Acyl-CoA dehydrogenases (ACADs), which are key enzymes in fatty acid and amino acid catabolism, form a large, pan-taxonomic protein family with at least 13 distinct subfamilies. Yet most reported ACAD members have no subfamily assigned, and little is known about the taxonomic distribution and evolution of the subfamilies. In completely sequenced genomes from approximately 210 species (eukaryotes, bacteria and archaea), we detect ACAD subfamilies by rigorous ortholog identification combining sequence similarity search with phylogeny. We then construct taxonomic subfamily-distribution profiles and build phylogenetic trees with orthologous proteins. Subfamily profiles provide unparalleled insight into the organisms' energy sources based on genome sequence alone and further predict enzyme substrate specificity, thus generating explicit working hypotheses for targeted biochemical experimentation. Eukaryotic ACAD subfamilies are traditionally considered as mitochondrial proteins, but we found evidence that in fungi one subfamily is located in peroxisomes and participates in a distinct beta-oxidation pathway. Finally, we discern horizontal transfer, duplication, loss and secondary acquisition of ACAD genes during evolution of this family. Through these unorthodox expansion strategies, the ACAD family is proficient in utilizing a large range of fatty acids and amino acids-strategies that could have shaped the evolutionary history of many other ancient protein families.

Figure 1.

Optimal substrates of ACAD subfamilies. C4, etc., length of the acyl-CoA chain. C16:1, unsaturated fatty acid with one double bond. Subfamilies in the left part of the figure are involved in fatty acid degradation. Those in the right part are involved in amino acid degradation. ‘R’ represents straight alkyl chain.

Figure 2.

Flow chart of the procedure for assigning ACAD subfamilies. Blast searches combined with reconciliation of gene and species trees were used to identify orthologs (see ‘Materials and Methods’ section). Y, yes; N, no.

Figure 3.

Distinction of ACD10 and ACD11. Protein domains of human ACD11 and ACD10 (Q709F0, A) and ACD10 (Q6JQN1, B). InterProt domain IDs are as follows: hydrolase domain, IPR005834; APH domain, IPR002575. The ACAD domain is composed of three parts: ACAD N-terminal domain, IPR013786; ACAD central domain, IPR006091; ACAD C-terminal domain, IPR013764. (C) Domain content of ACD10, ACD11 and provisional ACD10/11 homologs mapped onto the phylogenetic tree. Taxa representing more than three species are shown in bold. Clades with bootstrap support value >90 are labeled with asterisk. Taxa that appear twice in the tree are distinguished by the labels ‘ACD10’ and ‘ACD11’. In animals, ACD11 includes (in addition to the common ACAD domains) an APH domain, and ACD10 possesses an APH plus a hydrolase (Hyd) domain. Exceptions are gi|115941654 of the echinoderm Strongylocentrotus purpuratus, which is more similar to ACD10 but lack the hydrolase domain, and jgi|Dappu1|346313 of the crustacean Daphnia pulex, which shares equal sequence similarity with ACD10 and ACD11 and lacks both extra domains. Homologs of other eukaryotes, which have an APH domain, but no hydrolase domain, are classified as ACD11. Sequences lacking both domains are all homologs of fungi, the green algae Volvox carteri and Ostreococcus lucimarinus and the stramenopiles Aureococcus anophagefferens and Phytophthora ramorum. Bacterial homologs also lack both domains. Those lacking both domains are classified as ACD11n, see text.

Figure 4.

ACAD subfamily distribution mapped on the taxonomy hierarchy from NCBI. Only species whose genome has been completely sequenced are included in the figure. The sequence IDs are listed in ←Table S1. A triangle in front of a taxon name indicates that no ACAD subfamily was detected in the members of this taxon. (A) Subfamily distribution in prokaryotes; (B) subfamily distribution in eukaryotes.

Figure 4.

ACAD subfamily distribution mapped on the taxonomy hierarchy from NCBI. Only species whose genome has been completely sequenced are included in the figure. The sequence IDs are listed in ←Table S1. A triangle in front of a taxon name indicates that no ACAD subfamily was detected in the members of this taxon. (A) Subfamily distribution in prokaryotes; (B) subfamily distribution in eukaryotes.

Figure 5.

Alignment of residues that affect substrate specificity of human ACDSB. The multiple protein alignment was generated using eukaryotic ACDSB homologs. The numbers refer to the residue position of the mature human ACDSB (i.e. not including the mitochondrial targeting peptide). A minor deviation from the NIT motif is found in Puccinia graminis, which has a ‘NIS’.

Figure 6.

Schematic phylogenetic trees of ACAD subfamilies. The underlying explicit trees are provided in Figure S1. Branches with a bootstrap value ≥90 are labeled with filled dots. Taxa representing more than three species are shown in bold. (A) ACAD subfamilies with likely α-proteobacterial origin. In the trees of GCDH (left, Figure S1C) and IBD (right, Figure S1E), eukaryotic homologs group together with those from α-Proteobacteria. In the GCDH tree, eukaryotic and numerous α-proteobacterial taxa form a well-supported clade to the exclusion of Archaea plus Actinobacteria; both clusters include a few other Proteobacteria. The IBD tree unites eukaryotes and α-Proteobacteria to the exclusion of a few γ-Proteobacteria. (B) Secondary acquisition of α-proteobacterial homologs by certain fungal lineages. Some Basidiomycota and all investigated Ascomycota lack ACADM (Figure S1H). The ascomycete class Pezizomycotina possesses fadE12 (of same function as ACADM, Figure S1I) that associates strongly with α-Proteobacteria. Exceptions are Emericella nidulans, M. grisea and Chaetomium globosum, where fadE12 is absent. Ustilago maydis, the single basidiomycete possessing fadE12, likely acquired this gene from Pezizomycotina. (C) Gene duplication in animals and lateral transfer to other taxa. ACADV and ACADV2 are paralogs originating from a gene duplication prior to the divergence of animals (Figure S1O). The few bacterial ACADV homologs form a monophyletic clade, to the exclusion of animal proteins. ACADV2 from Phytophthora groups with animal homologs.