The Tsp45I restriction-modification system is plasmid-borne within its thermophilic host.
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.
Wayne
J
J
New England Biolabs Inc., Beverly, MA 01915, USA.
Holden
M
M
Xu
S Y
SY
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
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