STRUCTURE AND ORGANIZATION OF YEAST MAL LOCI

拟议的研究旨在研究分子 控制结构基因表达的机制 参与酵母中麦芽糖（和淀粉）的代谢。 我们 特别感兴趣的是， MAL基因座的调节成分参与诱导 麦芽糖和葡萄糖对碳分解代谢物的抑制。努力是 针对麦芽糖酶和麦芽糖的结构和功能， 通透酶启动子元件和反式作用调节元件的相互作用 蛋白质及其作用位点。 1. 完成MAL 6基因座的测序， MALR重复序列（MALRR）和MAL 1序列 基因间区（IG）。 在这些实验中， 整个MAL 6基因座和重复序列（MAL 6 RR） 将完成与MAL 6 R同源的研究。 的初步研究 MAL 6 RR的属性将被启动。 2. 关于MAL 6 IG中启动子元件的其他研究 地区 UASM序列将进行更详细的研究， 无论其方向和距离 TATA序列的重要性，因为它可能在 调节分化转录的通透酶基因。 的作用 葡萄糖阻遏中富含T序列将被进一步研究 并检测是否有特定的蛋白质与之结合。 3. MAL 6 R编码调控蛋白的研究 的 调节蛋白将在E.在大肠杆菌中合成， 为了制造针对它的抗体。调节蛋白 以及它的一些变异衍生物将被研究 用于与麦芽糖酶启动子区域相互作用。 4. 麦芽糖酶组成型和葡萄糖阻遏的研究 不敏感的突变体 组成型突变体将通过 诱变调节基因和IG区。 他们将 检查葡萄糖对它们的抑制作用。 5. MAL 1基因座的重复。 将构建二倍体 这将使人们能够分析随机孢子的复制品， MAL 1基因座。 6. 酵母糖化酶的研究。 STA 2基因的测序 将完成STA 10基因的克隆、STA基因的定位 并研究了MATa/MATalpha对STA基因的调控。

The proposed studies are designed to investigate the molecular mechanism(s) controlling the expression of the structural genes involved in maltose (and starch) metabolism in yeast. We are particularly interested in characterizing the cis and trans regulatory components of the MAL locus involved in induction by maltose and carbon catabolite repression by glucose. Effort is directed at the structure and function of the maltase and maltose permease promoter elements and of the trans-acting regulatory protein and its site(s) of action. 1. Complete the sequencing of the MAL6 locus, characterize the MALR repeated sequence (MALRR) and sequence the MAL1 intergenic (IG) region. In these experiments, the sequencing of the entire MAL6 locus and of a repeated sequence (MAL6RR) homologous to MAL6R will be completed. Preliminary studies on the properties of MAL6RR will be initiated. 2. Additional studies on the promoter elements in the MAL6 IG region. The UASM sequence will be studied in greater detail with respect to its boundaries, whether its orientation and distance from the TATA sequence are important and for its possible role in regulating the divergently transcribed permease gene. The role of the T-rich sequence in glucose repression will be studied further and examined to determine whether specific proteins bind to it. 3. Studies on the MAL6R encoded regulatory protein. The regulatory protein will be expressed in E. coli and synthesized in vitro in order to make antibodies to it. The regulatory protein and some of its mutationally altered derivatives will be studied for interaction with the maltase promoter region. 4. Search for maltase constitutive and glucose repression insensitive mutants. Constitutive mutants will be generated by mutagenizing the regulatory gene and the IG region. They will be checked for their repression by glucose. 5. Duplication of the MAL1 locus. Diploids will be constructed which will allow one to analyze random spores for duplications of the MAL1 locus. 6. Studies on yeast glucoamylase. Sequencing of the STA2 gene will be completed, the STA10 gene cloned, the STA genes mapped and the regulation of STA genes by MATa/MATalpha investigated.