| Project/Area Number |
62480057
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
発酵・醸造
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| Research Institution | HIROSHIMA UNIVERSITY |
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Principal Investigator |
MIYAKAWA Tokichi Hiroshima Univ. Fac. Engin., Prof., 工学部, 教授 (10116676)
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| Project Period (FY) |
1987 – 1989
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| Project Status |
Completed (Fiscal Year 1989)
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| Budget Amount *help |
¥6,800,000 (Direct Cost: ¥6,800,000)
Fiscal Year 1989: ¥700,000 (Direct Cost: ¥700,000)
Fiscal Year 1988: ¥1,000,000 (Direct Cost: ¥1,000,000)
Fiscal Year 1987: ¥5,100,000 (Direct Cost: ¥5,100,000)
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| Keywords | Yeast / Mating Pheromone / Farnesylation / クローニング / 遺伝子 |
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| Research Abstract |
Mating type A cells of the heterobasidiomycetous yeast Rhodosporidium toruloides, secrete mating pheromones rhodotorucine A that induce sexual differentiation in the cells of the opposite mating type (a). The pheromone is a S-farnesylated undecapeptide of quite interesting structure. To elucidate the mechanism of processing and secretion of the mating pheromone, we cloned and sequenced the genes for the mating pheromone. Important observations of our study are as follows: l.The genes for rhodotorucine A were cloned by screening genomic library of A cells using oligonucleotides synthesized according to the amino acid sequences of the mating pheromone as probe. From the nucleotide sequences of the clone, it was found that the deduced precursor protein for the mating pheromone contains four tandem repeats of the pheromone sequences, with a spacer sequence thr-val-ser-ala between the pheromone sequences. The spacer sequences may serve as a signal for the S-farnesylation of the peptide. Can
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onical signal sequences for protein secretion found at the N-terminus of almost all secretory proteins were not present in the precursor. From these results, we proposed that rhodotorucine A is secreted by a novel pathway in which modification of the peptide by the hydrophobic farnesyl residue plays an important role. Using immunoblot analysis, a polypeptide with the predicted molecular weight was found in the cell extract. 2.Three genes RHA1, RHA2 and RHA3 that encode four, three and five tandem repeats of rhodotorucine A sequences were found in the genome of A cells. 3.The RHA genes were found only in mating type A cells but not in a cells. The result was quite different from that known in the pheromone genes for Saccharomyces cerevisiar, in which the genes for mating pheromones are present in both mating types. 4.Oligopeptides that contain spacer sequences were synthesized according to the amino acid sequences of the precursor. Using the peptides and its analogues enzymatic reaction for the peetide farnesylation was measured. Less
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