The role of the Saccharomyces cerevisae peroxisomal acyl-coenzyme A (acyl-CoA) thioesterase (Pte1p) in fatty acid ␤-oxidation was studied by analyzing the in vitro kinetic activity of the purified protein as well as by measuring the carbon flux through the ␤-oxidation cycle in vivo using the synthesis of peroxisomal polyhydroxyalkanoate (PHA) from the polymerization of the 3-hydroxyacylCoAs 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 pte1⌬ 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 pte1⌬ strain compared with wild type. The poor catabolism of 8-methyl-nonanoic acid via ␤-oxidation in pte1⌬ negatively impacted the degradation of 10-cis-heptadecenoic acid and reduced the ability of the cells to efficiently grow in medium containing such fatty acids. An increase in the proportion of the short chain 3-hydroxyacid monomers was observed in PHA synthesized in pte1⌬ cells grown on a variety of fatty acids, indicating a reduction in the metabolism of short chain acyl-CoAs in these cells. A purified histidine-tagged Pte1p showed high activity toward short and medium chain length acyl-CoAs, including butyryl-CoA, decanoyl-CoA and 8-methyl-nonanoyl-CoA. The kinetic parameters measured for the purified Pte1p fit well with the implication of this enzyme in the efficient metabolism of short straight and branched chain fatty acyl-CoAs by the ␤-oxidation cycle.

A bioluminescent enzyme immunoassay (BLEIA) for prostate-specific antigen (PSA) using biotinylated firefly luciferase-labelled antibody was developed. PSA is an important marker for the diagnosis and management of prostate cancer. Our BLEIA for PSA, based on the two-step sandwich method, had ultra-high sensitivity and a very wide measurable range. The detection limit (mean of nine replicates of the zero standard +2 SD) for PSA was 0.25 pg/mL and the measurable range for PSA was 0.25 pg/mL-100 ng/mL. Generally, PSA in the serum exists on two forms, called free PSA (f-PSA) and complex PSA (c-PSA), which is formed with alpha-antichymotripsin. Thus, the response of the PSA assay to these two forms has to be equimolar in the construction of the assay system. Our BLEIA for PSA also had an equimolar response to them.

A 15-mer DNA labelled with a novel multifunctional fluorescent agent bearing acridine and fluorescein moieties is synthesized and the intensity of fluorescein-based fluorescence detected by the excitation of acridine is shown to be strongly affected by the DNA's secondary structure.

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. A specific feature of the methods is in a process of alternately feeding medium adjusted at two different nitrogen (N) concentrations. In the turbidostat-based method, a turbidostat culture was operated repeating three steps of determining D within a preset interval, alternating media by comparing the D with a preset value, and adjusting D in the next interval by feeding the selected medium. In the chemostat-based method, turbidity of a chemostat culture was controlled by repeating two steps of alternating media by comparing transmitted photon flux intensity (I) with a preset value and adjusting I by feeding the selected medium. D controlled by the turbidostat-based method reached quickly a preset value as low as 0.010/h, and then it was dispersed around but above the preset value. On the other hand, mean N concentrations of fed media formed a plateau. In the chemostat-based method, I was well controlled to a preset value while the mean N concentrations were a bit fluctuated. Starch concentration varied from 0.052 to 0.41 g/L with turbidity and D defined by these methods.