BIOCHEMISTRY & CYTOLOGY OF WALL SYNTHESIS IN FUNGI

这项研究的主要目的是了解甲壳素(几丁质)是如何 合成酶微囊)在体内工作，以及细胞 壁甲壳素微纤维是由一种正在生长的真菌合成的。我们以前的 研究表明，甲壳素合成酶的主要部分在 真菌以壳聚体的形式存在。这些微泡是 据信移动到细胞表面，在那里他们的几丁质合成酶酶原 被激活(可能是通过蛋白质分解)以及它们在哪里合成 甲壳素的确切位置和细节尚不清楚。一个 要研究的主要问题是壳聚糖体是否与 质膜，从而几丁质合成酶成为 这层膜。我们计划追踪细胞内的起源和命运 壳聚体及其与质膜相互作用的研究 体外培养的囊泡。为此，我们将努力开发关键的 分离最高纯度质膜样品的方法 正在生长的有壁的真菌细胞。我们将对两种真菌进行比较。此外 对于我们的主要测试生物体--鲁氏毛霉，我们还将检查脉孢子虫 Crassa，主要是因为它被证明适合制作质膜 研究。甲壳素合成酶的细胞定位将通过 无细胞萃取物的分级和通过金或 铁蛋白标记的抗甲壳素合成酶抗体。通过这些研究 我们希望解开真菌中几丁质原纤维聚集的机制。 墙壁。甲壳素不是毛霉中唯一由甲壳素合成酶合成的聚合物 Rouxii；我们发现该真菌通过一种协调的 糖基转移和脱乙酰化(通过甲壳素合成酶和甲壳素 脱乙酰酶。)这项研究的另一个主要目标是进一步 了解协调行动的细节，从而产生这一重要的 墙面聚合物。这项研究是一个长期项目的一部分，目的是阐明 毛霉形态发生的生化基础。

The main purpose of this study is to understand how chitosomes (chitin synthetase microvesicles) operate in vivo and the mechanism by which cell wall chitin microfibrils are synthesized by a growing fungus. Our previous studies have demonstrated that a major part of the chitin synthetase in fungi is present in the form of chitosomes. These microvesicles are believed to move to the cell surface where their chitin synthetase zymogen becomes activated (probably by proteolysis) and where they synthesize chitin by a mechanism whose precise site and details are still unknown. A major point to investigate is whether or not chitosomes fuse with the plasma membrane and thereby chitin synthetase becomes an integral part of this membrane. We plan to trace the intracellular origin and fate of chitosomes and to explore the interaction of chitosomes and plasma membrane vesicles in vitro. For this reason, we will endeavor to develop critical procedures to isolate plasma membrane samples of highest purity from growing, walled cells of fungi. Two fungi will be compared. In addition to Mucor rouxii, our main test organism, we will also examine Neurospora crassa, mainly because of its proven suitability for plasma membrane research. The cellular localization of chitin synthetase will be traced by fractionation of cell-free extracts and by means of gold-or ferritin-labelled anti-chitin synthetase antibodies. Through these studies we hope to unravel the mechanism of chitin fibril assembly in fungal walls. Chitin is not the only polymer made by chitin synthetase in Mucor rouxii; we have found that this fungus makes chitosan through a coordinated glycosyl transfer and deacetylation (via chitin synthetase and chitin deacetylase.) Another major goal of this study will be to further understand details of the coordinated action that produces this important wall polymer. This study is part of a long-standing program to elucidate the biochemical bases of morphogenesis of Mucor.