α-d-(1 → 3)-graft-(1 → 6)-glucan: Comb-like polysaccharide synthesized in vitro with α-1,3/1,6-glucosyltransferase L from Streptococcus salivarius.

We demonstrated that a unique polysaccharide with extremely high molecular weight can be easily obtained via a low-cost, mild reaction in a water medium from sucrose, a photosynthetic product. α-1,3/1,6-Glucosyltransferase L (GtfL) from Streptococcus salivarius produced water-insoluble α-d-glucan from sucrose at 37 °C. Gel permeation chromatography revealed the molecular weight was extremely high; the weight-average molecular weight values were more than 1,000,000 irrespective of the substrate concentration.

Biochemical and structural characterization of oxygen-sensitive 2-thiouridine synthesis catalyzed by an iron-sulfur protein TtuA.

Two-thiouridine (sU) at position 54 of transfer RNA (tRNA) is a posttranscriptional modification that enables thermophilic bacteria to survive in high-temperature environments. sU is produced by the combined action of two proteins, 2-thiouridine synthetase TtuA and 2-thiouridine synthesis sulfur carrier protein TtuB, which act as a sulfur (S) transfer enzyme and a ubiquitin-like S donor, respectively. Despite the accumulation of biochemical data in vivo, the enzymatic activity by TtuA/TtuB has rarely been observed in vitro, which has hindered examination of the molecular mechanism of S transfer.

Identification of a rhodanese-like protein involved in thiouridine biosynthesis in Thermus thermophilus tRNA.

Incorporation of a sulfur atom into 2-thioribothymidine (s T or 5-methyl-2-thiouridine) at position 54 in thermophile tRNA is accomplished by an elaborate system composed of many proteins which confers thermostability to the translation system. We identified ttuD (tRNA-two-thiouridine D) as a gene for the synthesis of s T54 in Thermus thermophilus. The rhodanese-like protein TtuD enhances the activity of cysteine desulfurases and receives the persulfide generated by cysteine desulfurases in vitro.

Folate-/FAD-dependent tRNA methyltransferase from Thermus thermophilus regulates other modifications in tRNA at low temperatures.

TrmFO is a N(5) , N(10) -methylenetetrahydrofolate (CH2 THF)-/FAD-dependent tRNA methyltransferase, which synthesizes 5-methyluridine at position 54 (m(5) U54) in tRNA. Thermus thermophilus is an extreme-thermophilic eubacterium, which grows in a wide range of temperatures (50-83 °C). In T.

Common thiolation mechanism in the biosynthesis of tRNA thiouridine and sulphur-containing cofactors.

2-Thioribothymidine (s(2)T), a modified uridine, is found at position 54 in transfer RNAs (tRNAs) from several thermophiles; s(2)T stabilizes the L-shaped structure of tRNA and is essential for growth at higher temperatures. Here, we identified an ATPase (tRNA-two-thiouridine C, TtuC) required for the 2-thiolation of s(2)T in Thermus thermophilus and examined in vitro s(2)T formation by TtuC and previously identified s(2)T-biosynthetic proteins (TtuA, TtuB, and cysteine desulphurases). The C-terminal glycine of TtuB is first activated as an acyl-adenylate by TtuC and then thiocarboxylated by cysteine desulphurases.