Yoshinari Yamazaki

Production and characteristics of some new β-agarases from a marine bacterium, Vibrio sp. strain JT0107

Abstract Vibrio sp. strain JT0107 is one of the marine bacteria that secrete β-agarases which catalyze the hydrolysis of agarose. The optimum culture conditions for the production of some β-agarases have been determined. To increase agarase activity, aeration and a sufficient concentration of agarose are needed. One of the enzymes that the bacteria secreted into the culture medium was isolated and purified 39-fold using a combination of ultrafiltration and subsequent anion exchange column chromatography. The purified protein migrated as a single band (72 kDa) on sodium dodecyl sulfate polyacrylamide gel electrophoresis and its isoelectric point was 4.7. Amino acid sequence analysis revealed a single N-terminal sequence that had no sequence identity to other marine bacterial agarases. This novel enzyme was found to be an endo-type β-agarase (EC 3.2.1.81) that catalyzes the hydrolysis of the β-1,4 linkage of agarose to yield neoagarotetraose [O-3,6-anhydro-α- l -galactopyranosyl(1→3)-O-β- d -galactopyranosyl(1→4)-O-3,6-anhydro-α- l -galactopyranosyl(1→3)- d -galactose] and neoagarobiose [O-3,6-anhydro-α- l -galactopyranosyl(1→3)- d -galactose]. The optimum pH and temperature for obtaining high activity of the enzyme were at around 8 and 30°C, respectively. The enzyme did not degrade sodium alginate, λ-carrageenan, ι-carrageenan or κ-carrageenan.

Synthesis of 5-methyluridine by a thermophile, Bacillus stearothermophilus JTS 859

Many thermophiles, which can grow at 65°C, were examined as to their ability to produce 5-methyluridine from inosine and thymine (5-methyluracil) in the presence of phosphate and cells as enzyme sources. Bacillus stearothermophilus JTS 859 was selected as a strain that synthesized 5-methyluridine efficiently. The reaction is supposed to be carried out by a combination of a thermostable purine nucleoside phosphorylase and a thermostable pyrimidine nucleoside phosphorylase. Their half-lives were 7200 hr and 400 hr at 63°C, and 148 hr and 14 hr at 70°C, respectively.