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Table 1 Summary of features of the Thermus sp.enzyme family

From: Related bifunctional restriction endonuclease-methyltransferase triplets: TspDTI, Tth111II/TthHB27I and TsoI with distinct specificities

REase TspDTI Tth111II TthHB27I TsoI TspGWI TaqII
Bacterial host Thermus sp. DT Thermus thermophilus 111 Thermus thermophilus HB27 Thermus scotoductus Thermus sp. GW Thermus aquaticus YTI
Recognition and cleavage site ATGAA(11/9) CAARCA(11/9) CAARCA(11/9) TARCCA(11/9) ACGGA(11/9) GACCGA(11/9)
Recognition site length (bp) 5 6, (degenerated) 6 (degenerated) 6 (degenerated) 5 6
ORF GC% 57.99 61.67 61.59 56.54 69.19 66.27
Polypeptide length (aa) 1112 1106 1121 1116 1097 1106
Molecular weight (kDa) 126.885 125.956 127.672 126.474 120.202 125.675
Native molecular size (kDa)/protein organization 110-130 monomer 110-130 monomer ND ND 110-130 monomer 110-130 monomer
Optimum reaction temperature 65°C 65-70°C ND 55°C 65°C 65°C
Single site cleavage Yes ND ND ND Two sites preferred Yes
Isoelectric point (calculated) 6.68 6.60 6.80 8.11 6.58 5.40
Specific DNA + methylation   ND ND + + +
Cleavage stimulation with SAM Yes ND ND Yes Slight inhibition Yes
Cleavage stimulation with SIN (SAM analogue) Yes ND ND ND Yes, specificity change Yes, specificity change
Methylation stimulation with Ca2+ No ND ND ND Yes Yes
Cleavage stimulation with divalent cations Ca2+ ND ND ND Mn2+, Fe2+, Co2+ not stimulated by Ca2+ ND
Domain organization* R-Mh-Mc-S R-Mh-Mc-S R-Mh-Mc-S R-Mh-Mc-S R-Mh-Mc-S R-Mh-Mc-S
Cleavage catalytic motif (B/E)** atypical
atypical D-EXE (B) atypical D-EXE (B) atypical D-EXE (B) PDX13EX1K (B + E) PDX13EX1K (B + E)
Methylation catalytic motif (B/E)** NPPW (B) NPPW (B) NPPW (B) NPPW (B) NPPY (B + E) NPPY (B + E)
Subfamily homology* Subfamily TspDTI Subfamily TspDTI Subfamily TspDTI Subfamily TspDTI Subfamily TspGWI Subfamily TspGWI
Reference [20] GenBank: AY726624.1, [20, 26] GenBank: AE017221.1 This work, manuscript in preparation [19, 23] GenBank: AY057443, 21
  1. * Domain organization: R, REase catalytic domain; Mh, MTase helical domain; Mc, MTase catalytic domain; S, putative specific DNA recognition domain
  2. ** B, determined by bioinformatics analysis; E, experimentally determined by site-directed mutagenesis.