Genetic replacement of tesB with PTE1 affects chain-length proportions of 3-hydroxyalkanoic acids produced through β-oxidation of oleic acid in Escherichia coli.

Acyl-CoA thioesterase II (TesB), which catalyzes hydrolysis of acyl-CoAs to free fatty acids and CoA, is involved in 3-hydroxyalkanoic acid production in Escherichia coli. Effects of genetic replacement of tesB with Saccharomyces cerevisiae acyl-CoA thioesterase gene PTE1 on 3-hydroxyalkanoic acid production from oleic acid through β-oxidation were examined. Kinetic analyses using β-oxidation intermediates showed that hydrolyses of C4-acyl substrates are more efficient by PTE1 than by TesB.

The peroxisomal Acyl-CoA thioesterase Pte1p from Saccharomyces cerevisiae is required for efficient degradation of short straight chain and branched chain fatty acids.

The role of the Saccharomyces cerevisae peroxisomal acyl-coenzyme A (acyl-CoA) thioesterase (Pte1p) in fatty acid beta-oxidation was studied by analyzing the in vitro kinetic activity of the purified protein as well as by measuring the carbon flux through the beta-oxidation cycle in vivo using the synthesis of peroxisomal polyhydroxyalkanoate (PHA) from the polymerization of the 3-hydroxyacyl-CoAs as a marker. The amount of PHA synthesized from the degradation of 10-cis-heptadecenoic, tridecanoic, undecanoic, or nonanoic acids was equivalent or slightly reduced in the pte1Delta strain compared with wild type. In contrast, a strong reduction in PHA synthesized from heptanoic acid and 8-methyl-nonanoic acid was observed for the pte1Delta strain compared with wild type.

Simultaneous control of turbidity and dilution rate through adjustment of medium composition in semi-continuous Chlamydomonas cultures.

For production of starch in algal cultures, a growth rate limited by a nutrient is an important factor. Under phototrophic conditions, turbidity must be also paid attention, as the shading effect may affect its productivity. Semi-continuous cultivation methods, which enable control of turbidity and dilution rate (D) at the same time, have been developed for evaluation of those factors on starch production in Chlamydomonas sp.

Preparation of a conjugate of 2',5'-triadenylate 5'-triphosphate and biotinylated firefly luciferase and its use for sensitive bioluminescent enzyme immunoassay.

A bioluminescent enzyme immunoassay (BLEIA) for 2',5'-oligoadenylate 5'-triphosphate (pppA2'p5'A2'Ap5'A, 2-5A) was developed using a conjugate of 2-5A and firefly luciferase as a probe. The conjugate was prepared from a 2-5A derivative bearing a thiol group at the 2'(or 3') end, and streptavidin (SA) bearing maleimide via a linker-arm and biotinylated luciferase. The thiol group of the 2-5A derivative was relatively stable under aerobic conditions and remained 100% and 56% intact at 25 degrees C after 1 and 96 h, respectively, without dimerization by aerobic oxidation.