title: The Tsp45I restriction-modification system is plasmid-borne within its thermophilic host.
abstract: Thermus species YS45 harbors two small cryptic plasmids of 5.8 (pTsp45s) and approximately 12 kb (pTsp45I). Plasmid pTsp45s has been entirely sequenced, revealing three significant ORFs. In addition to a previously reported thermophilic plasmid-encoded replication protein (Rep), pTsp45s contains two genes for the Tsp45I methyltransferase (M.Tsp45I) and restriction endonuclease (Tsp45I). These two converging genes (tsp45IM and tsp45IR) overlap by 4 bp at their stop codons within an XbaI site. M.Tsp45I (413 aa, 47.0 kDa, recognizing 5'-GTSAC-3') is highly homologous to other m6A-methyltransferases, especially M.EcaI (recognizing 5'-GGTNACC-3'). Tsp45I (332 aa, 37.4 kDa, cleaving 5'-/GTSAC-3') is not homologous to M.Tsp45I, or to other restriction endonucleases. Recombinant Tsp45I is stably produced in E. coli, and cleaves DNA at 65 degrees C with the same specificity as the native enzyme. Therefore, the thermophilic Tsp45I restriction-modification system is plasmid-borne within its native host.
authors.affiliation: New England Biolabs Inc., Beverly, MA 01915, USA.
authors.forename: J
authors.initials: J
authors.lastname: Wayne
authors.forename: M
authors.initials: M
authors.lastname: Holden
authors.forename: S Y
authors.initials: SY
authors.lastname: Xu
mesh_terms: D000595:Amino Acid Sequence; D001426:Bacterial Proteins; D001483:Base Sequence; D015254:DNA Modification Methylases / Q000737:chemistry; D015252:Deoxyribonucleases, Type II Site-Specific / Q000737:chemistry / Q000235:genetics*; D008969:Molecular Sequence Data; D010957:Plasmids / Q000737:chemistry / Q000235:genetics*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific) / Q000737:chemistry / Q000235:genetics*; D013696:Temperature; D013824:Thermus / Q000201:enzymology* / Q000235:genetics* / Q000502:physiology
chemical_list: D001426:Bacterial Proteins; D015254:DNA Modification Methylases; C109784:Tsp45I methylase, Thermus sp. YS45; D015265:Site-Specific DNA-Methyltransferase (Adenine-Specific); C083548:Tsp45I endonuclease, Thermus sp. YS45; D015252:Deoxyribonucleases, Type II Site-Specific