Figure 2: Multiple amino acid sequence alignment of the closest TthPrimPol orthologues.
From: TruePrime is a novel method for whole-genome amplification from single cells based on TthPrimPol
The first block of sequences corresponds to Thermales (Thermus and Meiothermus) and the second block includes Si/pRN1 PrimPol and some other putative bifunctional primases/polymerases from bacteria, archaea and phage; in addition, plasmidic Rep (a potential PrimPol) and BcMCM PrimPol were included. Numbers in parentheses indicate the number of amino acid residues not shown. Invariant or conserved residues among Thermales (first set of sequences) were labelled in red and blue letters, respectively. Identity matches in the second set of sequences were equally coloured. The alignment defines several conserved regions, including the highly conserved motifs A, B and C (boxed in yellow), characteristic of AEP-like primases. Experimentally determined secondary structure elements in Si/pRN1 PrimPol are indicated above pRN1 sequence (α-helices, lettered cylinders; green) and β-strands (numbered arrows; orange). Modelled secondary structure elements in TthPrimPol are tentatively depicted above the TthPrimPol sequence (see also Fig 1b). The corresponding aligned regions are boxed in the same colours to emphasize structural conservation between TthPrimPol and the AEP core of pRN1. The C-terminal region of TthPrimPol, conserved in other Thermales (boxed in grey), aligns with the PriCT-1 domain of Rep and Eph primases; this region is not yet crystallized in pRN1 and it has been described as pRN1_helical29. Dots indicate invariant residues acting either as metal (red), nucleotide (purple) or Zn (magenta) ligands. Selection of the closest TthPrimPol homologues and multiple alignment of their amino acid sequence were initially performed with the BLAST programme and further adjusted manually to maximize similarities with the structured regions of pRN1 PrimPol. Nomenclature: YP_004631.1 T. thermophilus (Tth); YP_006971229.1 Thermus oshimai (Tos); WP_003046664.1 Thermus aquaticus (Taq); YP_004202830.1 Thermus scotoductus (Tsc1); YP_004202855.1 T. scotoductus (Tsc2); ETN89075.1 Thermus sp (Tsp); YP_003508539.1 Meiothermus ruber (Mru); YP_003684976.1 Meiothermus silvanus (Msi1); YP_003684747.1 M. silvanus (Msi2); AAC44111.1 plasmid pRN1 ORF904 from S. islandicus (pRN1); YP_502469.1 Methanospirillum hungatei (Mhu); YP_006262572.1 Deinococcus gobiensis (Dgo); YP_002829910.1 S. islandicus (Sis); YP_003357218.1 Methanocella paludícola (Mpa); AFO10831.1 Enterococcus phage EfaCPT1 (Eph); YP_009074444.1 Shigella sonnei Rep protein from plasmid ColE4-CT9 (Rep); WP_044797243 PrimPol-helicase from B. cereus (BcMCM).
