Thermostable RNA ligase from thermus phage
View Patent ↗An RNA ligase derived from bacteriophage TS2126 which infects Thermus scotoductus , nucleic acids comprising nucleotide sequences of open reading frame (ORF) and polypeptides encoded by the nucleic acids, are described.
1. An isolated polypeptide which is a RNA ligase selected from the group consisting of:
a) a polypeptide comprising the amino acid sequence of SEQ ID NO: 2;
b) a polypeptide encoded by a nucleic acid comprising the sequence of SEQ ID NO: 1; and
c) active fragments thereof.
2. The polypeptide of claim 1 , which is a fusion polypeptide.
3. An isolated thermostable RNA ligase having at least 95% sequence identity to the amino acid sequence of SEQ ID NO: 2.
4. The RNA ligase of claim 3 , wherein the ligase catalyzes ligation of single-stranded DNA, wherein ligation efficiency is measured using a single-stranded 22-mer DNA 5′P-d(N 22 ) as a substrate under the following conditions:
0.1 mg/mL ligase is reacted with 5 μM of said substrate in a solution with 1 mM ATP, 50 mM MOPS (pH 7.5), 5 mM MgCl 2 , 1 mM DTT, 10 mM KCl, and 25 μg/mL BSA, and
the ligase produces over 50% ligation of the single-stranded DNA after 2 hours at 60° C.