TAKATERA Kikuo University of Tokyo, Institute of Industrial Science, Technical associate, 生産技術研究所, 教務職員 (40216669)
YOSHIDA Shoichiro University of Tokyo, Institute of Industrial Science, Reserch associate, 生産技術研究所, 助手 (40111570)
|Budget Amount *help
¥1,700,000 (Direct Cost : ¥1,700,000)
Fiscal Year 1994 : ¥300,000 (Direct Cost : ¥300,000)
Fiscal Year 1993 : ¥1,400,000 (Direct Cost : ¥1,400,000)
(1) By combining reversed-phase high-performance liquid chromatography (RP-HPLC) with inductively coupled plasma mass spectrometry (ICPMS), the induction of two major metallothionein (MT) isoforms has been demonstrated for a cyanobacterium Anacystis nidulans R-2, cultured under a stress of Zn^<2+> or Cd^<2+>. The two isoforms showed only a slight difference in the amino acid composition. Incubation of two Zn-MT isoforms in a medium containing Cd^<2+> led to a metal exchange, reflecting the higher affinity of MT to Cd^<2+> than to Zn^<2+>. During this process the relative abundance of the two isoforms remained practically unchanged. The two isoforms were induced in a constant ratio by the Zn^<2+> dosage (1-20 muM), while by the Cd^<2+> dosage (0.5-10 muM) the ratio showed a change with the dose level. These results demonstrate the potential of the present RP-HPLC/ICPMS system to elucidate the function of MT isoforms.
(2) Selenium is a toxic element which can cause environmental pollution
. Its uptake and incorporation by Zn^<2+>-induced metallothionein (MT) biosynthesis in a cyanobacterium were studied by means of HPLC/ICPMS.When we cultured the cells in selenite-enriched media, as muchas 60% of the total selenium in the water-soluble cell components was incorporated into MT,which exists as two major isoforms but occupies a very minor fraction of the overall proteins. Existence of two pathways has been shown for the first time for selenium incorporation into the cyanobacterial MT ; one is the S-Se bonding which can be reductively broken by 2-mercaptoethanol, and the other is the formation of selenocysteine followed by its incorporation into the MT peptide chain. No essential difference was noted between selenite and selenate regarding the uptake or the incorporation of Se. The present HPLC/ICPMS system can elucidate biological processes of environmental chemical importance at the molecular level.