Table 2 A list of active BisI homolog enzymes that prefer to cut GCNGC sites with three to four m5C.
REase | GenBank accession # or locus tag | Bacterial strain | # of aa & aa seq identity to BisI b | Activity on pBRFM (2–3 m5C) | Cut sites & Comment c |
|---|---|---|---|---|---|
Eco15I | EIL45146 | E. coli 541-15 | 290 aa (26.2%, N-terminal 160 aa) | Yes | 3–4 m5C > 2 m5C |
Sde240I (PK7) | WP_041325806 | Saccharophagus degradans | 162 aa (24.3%) | Yes | 3–4 m5C > 2 m5C |
Pan13I | WP_014606222 | Pantoea ananatis PA13 | 157 aa (24.0%) | Yes | 3–4 m5C > 2 m5C |
Pru4541I | EFB72318 | Providencia rustigianii DSM 4541 | 173 aa (23.9%) | Yes (3 m5C sites) | 3–4 m5C > 2 m5C |
AlaI (PK4) | WP_026374325 | Agrococcus lahaulensis | 160 aa (23.0%) | Yes | 3–4 m5C > 2 m5C |
Sve396I | EFQ58914 | Streptococcus vestibularis F0396 | 164 aa (22.3%) | Yes (3 m5C sites) | 3–4 m5C |
SqiI a (Pk9) | WP_026758869 | Sediminimonas qiaohouensis | 161 aa (22.3%) | No | 4 m5C |
Dsp20IU (PK5) | KDB03925 | Defluviimonas sp. 20V17 | 161 aa (22.1%) | Yes | 3–4 m5C > 2 m5C. prefer GCWGC |
NhoI | CCF83679 | Nitrolancetus hollandicus Lb | 169 aa (21.9%) | Yes (3 m5C sites) | 3–4 m5C |
SmaAUI | EMI48892 | Stenotrophomonas maltophilia AU12-09 | 168 aa (20.4%) | Yes | 3–4 m5C > 2 m5C |
MbaR4I (PK3) | WP_017202926 | Microbacterium barkeri | 190 aa (17.2%) | Yes | 3–4 m5C > 2 m5C |
2 m5C
3 m5C
2 m5C- aSqiI requires four m5C in GCWGC (4 m5C
3 m5C) for activity. The cut site of SqiI and MbaR4I was inferred from digestion of modified duplex oligos. - bPairwise sequence alignment by EMBOSS Needle server (gap penalty = 10.0, extend penalty = 0.5). List of enzymes in descending order of aa sequence identity to BisI.
- cCut sites were determined by run-off sequencing of enzyme-digested pBRFM substrate. The cleavage efficiency of the modified site (3–4 m5C) was carried out on duplex oligos. The cut sites of Eco15I, Pan13I, Pru4541I, SmaAUI, Sve396I, and NhoI were determined by run-off sequencing of enzyme-digested pBRFM substrate (GCNGC sites with two or three m5C).
3 m5C) for activity. The cut site of SqiI and MbaR4I was inferred from digestion of modified duplex oligos.