Evolutionary relationships of ATP-Binding Cassette (ABC) uptake porters

Abstract

Background: The ATP-Binding Cassette (ABC) functional superfamily includes integral transmembrane exporters that have evolved three times independently, forming three families termed ABC1, ABC2 and ABC3, upon which monophyletic ATPases have been superimposed for energy-coupling purposes [e.g., J Membr Biol 231(1):1-10, 2009]. The goal of the work reported in this communication was to understand how the integral membrane constituents of ABC uptake transporters with different numbers of predicted or established transmembrane segments (TMSs) evolved. In a few cases, high resolution 3-dimensional structures were available, and in these cases, their structures plus primary sequence analyses allowed us to predict evolutionary pathways of origin.

Results: All of the 35 currently recognized families of ABC uptake proteins except for one (family 21) were shown to be homologous using quantitative statistical methods. These methods involved using established programs that compare native protein sequences with each other, after having compared each sequence with thousands of its own shuffled sequences, to gain evidence for homology. Topological analyses suggested that these porters contain numbers of TMSs ranging from four or five to twenty. Intragenic duplication events occurred multiple times during the evolution of these porters. They originated from a simple primordial protein containing 3 TMSs which duplicated to 6 TMSs, and then produced porters of the various topologies via insertions, deletions and further duplications. Except for family 21 which proved to be related to ABC1 exporters, they are all related to members of the previously identified ABC2 exporter family. Duplications that occurred in addition to the primordial 3 → 6 duplication included 5 → 10, 6 → 12 and 10 → 20 TMSs. In one case, protein topologies were uncertain as different programs gave discrepant predictions. It could not be concluded with certainty whether a 4 TMS ancestral protein or a 5 TMS ancestral protein duplicated to give an 8 or a 10 TMS protein. Evidence is presented suggesting but not proving that the 2TMS repeat unit in ABC1 porters derived from the two central TMSs of ABC2 porters. These results provide structural information and plausible evolutionary pathways for the appearance of most integral membrane constituents of ABC uptake transport systems.

Conclusions: Almost all integral membrane uptake porters of the ABC superfamily belong to the ABC2 family, previously established for exporters. Most of these proteins can have 5, 6, 10, 12 or 20 TMSs per polypeptide chain. Evolutionary pathways for their appearance are proposed.

Figure 1

Internal 3 TMS repeats in 6 TMS proteins. A (left). Hydropathy plot of MalG (TC# 3.A.1.1.1), a six TMS membrane porter. Blue lines denote Hydropathy; Red lines denote Amphipathicity; Orange bars mark transmembrane segments as predicted by HMMTOP. B (right). TMSs 1–3 of gi220933130 aligning with TMSs 4–6 of gi255331744 yielded a comparison score of 10.9 S.D. with 40.3% similarity and 27.7% identity. The numbers at the beginning of each line refer to the residue numbers in each of the proteins. TMSs are indicated in red lettering. Vertical lines indicate identities; colons indicate close similarities, and periods indicate more distant similarities.

Figure 2

Internal 6 TMS repeats in 12 TMS proteins. A (left). Hydropathy plot of the ferric iron porter, FutB. Blue lines denote hydropathy; Red lines denote amphipathicity; Orange bars mark transmembrane segments as predicted by HMMTOP. B (right). TMSs 7– 12 of gi163796270 aligned with TMSs 1–6 of gi113476753, yielding a comparison score of 13.7 S.D. with 36.3% similarity and 27.1% identity. The numbers at the beginning of each line refer to the residue numbers in each of the proteins. TMSs are indicated in red lettering. Vertical lines indicate identities; colons indicate close similarities, and periods indicate more distant similarities.

Figure 3

Internal 5 TMS repeats in some 10 TMS transporters. A (left). Hydropathy plot of RnsC (TC# 3.A.1.2.12). Blue lines denote Hydropathy; Red lines denote Amphipathicity; Orange bars mark transmembrane segments as predicted by HMMTOP. B (right). Putative TMSs 1– 5 of gi222147212 are aligned with putative TMSs 6–10 of gi218884703, yielding a comparison score of 14.9 S.D. with 41.1% similarity and 29.5% identity. The numbers at the beginning of each line refer to the residue numbers in each of the proteins. TMSs are indicated in red lettering. Vertical lines indicate identities; colons indicate close similarities, and periods indicate more distant similarities.

Figure 4

Hydropathy plot of the BtuC (TC# 3.A.1.13.1) vitamin B₁₂ transport protein. The topological prediction was performed with the WHAT program. Blue lines denote Hydropathy; Red lines denote Amphipathicity; Orange bars mark transmembrane segments as predicted by HMMTOP.

Figure 5

Red lettering indicates the TMSs (TM1-10) as also indicated by the helical structures above the sequence. Numbers at the beginning of each line refer to the residue numbers in the protein. TMSs within BtuC revealed by x-ray crystallography.

Figure 6

TMSs 1–4 of gi288941543 aligned with TMSs 6–10 of gi150017008, giving a comparison score of 13.6 S.D. with 42.1% similarity and 31.0% identity. The numbers at the beginning of each line refer to the residue numbers in each of the proteins. TMSs are indicated in red lettering. Vertical lines indicate identities; colons indicate close similarities, and periods indicate more distant similarities.

Figure 7

TMSs 1–3 of gi108803469 aligned with TMSs 1–3 of gi126656877. The comparison score was 11.2 S.D. with 42.6% similarity and 30.9% identity. The numbers at the beginning of each line refer to the residue numbers in each of the proteins. TMSs are indicated in red lettering. Vertical lines indicate identities; colons indicate close similarities, and periods indicate more distant similarities.

Figure 8

TMSs 5–7 of gi295100997 aligning with TMSs 4–6 of gi13471902. The comparison score was 11 S.D. with 32.5% similarity and 20.1% identity. The numbers at the beginning of each line refer to the residue numbers in each of the proteins. TMSs are indicated in red lettering. Vertical lines indicate identities; colons indicate close similarities, and periods indicate more distant similarities.

Figure 9

Putative TMSs 1–4 of an RnsC homologue (gi31544792) (top) aligned with putative TMSs 1–4 of the six TMS MalG homologue (gi116512192) (bottom). The comparison shown was 12.7 S.D. (45% similarity and 22.5% identity). The numbers at the beginning of each line refer to the residue numbers in each of the proteins. TMSs are indicated in red lettering. Vertical lines indicate identities; colons indicate close similarities, and periods indicate more distant similarities.

Figure 10

TMSs 1–3 compared with TMSs 4–6 of an ABC type 2 ancestral sequence. The comparison score was 39.9 SD with 58.5% similarity and 50.4% identity. The numbers at the beginning of each line refer to the residue numbers in each of the proteins. TMSs are indicated in red lettering. Vertical lines indicate identities; colons indicate close similarities, and periods indicate more distant similarities.

Figure 11

Structural superimposition of MalF and MalG. A (left). The last 3 TMS domain-duplicated unit of MalF (TMSs 6, 7 and 8) superposed on that of MalG (TMSs 4, 5 and 6). The TMS numbering shown is taken from MalG. The light colored chain represents MalG, and the coordinates used are the X/Y coordinate columns. B (right). The first 3 TMS domain-duplicated unit of MalF (TMSs 3, 4 and 5) superposed on the first duplicated unit of MalG (TMSs 1, 2 and 3). The TMS numbering shown is for MalF. The light colored chain represents MalF, and the coordinates used are the Y/Z coordinate columns.

Figure 12

Possible origin of ABC1 porters from ABC2 porters. TMSs 3 and 4 of an ABC1 homologue, gi283948596 (top), aligned with TMSs 3 and 4 of an ABC2 homologue, gi149372921 (bottom), giving a comparison score of 11 S.D, 52.5% similarity and 39% identity. The numbers at the beginning of each line refer to the residue numbers in each of the proteins. TMSs are indicated in red lettering. Vertical lines indicate identities; colons indicate close similarities, and periods indicate more distant similarities.

Figure 13

Cytoscape 2.8.3 graph, using spring embedded logic, of significant relationships between all families within 3.A.1.

Figure 14

Proposed evolutionary pathway and primordial sequences of the different topological types of ABC uptake systems. A (left). The proposed evolutionary pathway for the appearance of present-day ABC uptake systems. B (right). Presumed primordial or intermediate sequences and representative examples of the different topological types of ABC transmembrane porter proteins.