9405844 Matthysse It was propose to continue our investigation of the mechanisms of cellulose synthesis by A. tumefaciens. In previous research, two operons required for cellulose synthesis were identified and sequenced. In the same studies, an in vitro cellulose synthesis system was also developed. With this in vitro cellulose synthesis system together with mutant cells in the cel (cellulose synthesis) genes, the role of each one of the cel gene products in cellulose formation may be determined. By using the same in vitro cellulose synthesis system and mutant collection, putative intermediates in cellulose synthesis were found. The chemical structure of these intermediates will be determined. In addition, cellulose synthesis in A. tumefaciens is increased in the present of plant cells or plant extracts. The mechanism of this regulation will be examined. %%% Cellulose, which is made by plants and by some bacteria and fungi, is one of the most abundant biologically produced materials on earth. It is a relatively simple molecule consisting of a linear chain of beta-1,4-glucose molecules. However, the detailed mechanism of cellulose synthesis is not well understood. It is not now known whether all organisms which make cellulose use similar pathways, or whether there are different pathways used by different groups of organisms. The regulation of cellulose synthesis is also poorly understood. In this research project, the mechanism and regulation of cellulose synthesis by a bacterium called Agrobacterium tumefaciens will be examined. The advantages of using a bacterium to study cellulose synthesis are that the bacteria are relatively simple and often allow more easily for the determination of genes required and the individual role(s) of the same gene products in a complex biosynthetic pathway. In an earlier study, the genes for cellulose formation by this bacterium were determined. Now with a collection of mutant cells and the development of a cell-fre e cellulose synthesis system, it is hoped that the function of each one of the genes of cellulose synthesis can be determined, that the chemical steps in the pathway for cellulose formation will be identified, and that experiments can be undertaken to begin to understand how the microbial pathway is regulated for cellulose formation. ***
