Studies on the biosynthesis and secretion of mating pheromone (farnesyl peptide) in a heterobasidiomycetous yeast.

异形担子酵母的交配型A型细胞分泌交配信息素Rhodotorucine A，在相反交配型(A)的细胞中诱导性分化。信息素是一种S法尼化的十一肽，结构非常有趣。为了阐明交配信息素的加工和分泌机制，我们对交配信息素基因进行了克隆和测序。1.根据交配信息素的氨基酸序列合成的寡核苷酸作为探针，通过筛选A细胞基因组文库，克隆了红曲霉毒素A的基因。从克隆的核苷酸序列中发现，推测的交配信息素前体蛋白含有信息素序列的四个串联重复序列，信息素序列之间有一个间隔序列thr-val-ser-ala。间隔区序列可作为多肽S法尼化的信号。能否…在几乎所有分泌蛋白的N端发现的更多分泌蛋白质的信号序列在前体中不存在。根据这些结果，我们认为柔红曲霉素A是通过一种新的途径分泌的，其中疏水的法尼基残基对多肽的修饰起着重要作用。通过免疫印迹分析，在细胞提取液中发现了一种具有预测分子量的多肽。2.在A细胞基因组中发现了三个基因RHA1、RHA2和RHA3，它们编码四个、三个和五个Rhodotorucine A序列的串联重复序列。3.RHA基因仅在交配型A细胞中发现，而在A细胞中未发现。这一结果与酿酒酵母信息素基因的已知结果大不相同，在酿酒酵母信息素基因中，交配信息素基因在两种交配类型中都存在。4.根据前体的氨基酸序列合成了含有间隔区序列的寡肽。利用所合成的多肽及其类似物进行了多肽法尼化反应的测定。较少

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 … More 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