| Budget Amount *help |
¥2,100,000 (Direct Cost: ¥2,100,000)
Fiscal Year 1989: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1988: ¥1,400,000 (Direct Cost: ¥1,400,000)
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| Research Abstract |
ATP synthase (CF^0 CF^1) of chloroplast catalyzes synthesis of ATP coupled to an electrochemical gradient of protons. The catalytic portion, CF^1, consists of five different subunits alpha, beta, gamma, delta, and epsilon. Energization of chloroplast thylakoids activates CF^0 CF^1, to catalyze ATP synthesis and ATP hydrolysis. In the activation processes, CF^0 CF^1, seems to be in several distinct states. This suggests distinct conformations of CF^0 CF^1, in the activation processes.
To know the distinctions among the conformations of CF^0 CF^1, it is useful to map the amino acid residues which are under the influence of the changes in conformation of CF^0 CF^1. If the surroundings of one amino acid residue change greatly accompanying the changes in conformation of CF^0 CF^1, the reactivity of the amino acid residue is expected to alter. In this report, the changes in reactivity of lysine residues in the epsilon subunit of CF^1 were studied by using pyridoxal 5'-phosphate.
Incubation of spinach chloroplast thylakoids with pyridoxal 5'-phosphate modified the epsilon subunit of ATP synthase CF^0 CF^1. Illumination of thylakoids stimulated the modification of one specific amino acid residue of the epsilon subunit by a factor of three. Endoproteinase Glu-C treatment of the isolated epsilon subunit and fractionation of the peptides by high performance liquid chromatography revealed a major fluorescent peptide with a sequence GKRQKIE. Further treatment of this peptide with Endoproteinase Arg-C gave a strongly fluorescent tripeptide GXR. From the primary structure of the epsilon subunit, the specifically modified residue was deduced to be Lys-109.
This suggests the energy-dependent conformational changes in the epsilon subunit which change the surroundings of Lys-109 and alter the reactivity of this residue.
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