2002 Annual Report 1.What major problem or issue is being resolved and how are you resolving it? The successful production of cereals with value-added traits depends upon our ability to consistently obtain optimal expression of the desired traits. We cannot yet consistently produce cereal grains that meet the stringent specifications required by the processing industries that use the grain. In part, this is due to an incomplete knowledge of the physiology and biochemistry of cereals. Additional limitations result from the ill-defined and changing needs of specific industries as well as imprecise measurements of these qualities. We are working to complete our understanding of the biochemical and physiological processes that result in the conversion of starch to sugars, which is a crucial value-added trait for malting barleys. Research from this program will potentially lead to development of new or improved crop products, and it will define the properties of value-added traits in such a way that better measurements of quality result. New, more accurate measurements of malt quality will provide the basis for improved pricing of barleys shown to have high malting quality for customers. 2.How serious is the problem? Why does it matter? Growers who lose an anticipated premium (typically $0.25-1.00/bushel) for malting quality barley consider crop rejection by malting/brewing industries to be a significant loss. The problem is quite serious to industrial users as unexpected variation in the raw product (malt) requires additional expenses in downstream processing. 3.How does it relate to the national Program(s) and National Program Component(s) to which it has been assigned? This research contributes primarily to solving problem areas IIa and IIc of National Program 302 by providing fundamental and applied information on a value-added trait critical to industry and to the growers/producers of the raw material who receive the market premium for the crop. 4.What was your most significant accomplishment this past year? A. Single Most Significant Accomplishment during FY2002: Demonstrated the significance of including thermostable alpha-glucosidase, produced via site directed mutagenesis, during mashing upon the carbohydrate profile and the real degree of fermentation values (RDF) obtained in the final wort. The RDF values of mashes containing thermostable enzyme were 1.7 times greater per unit of added enzyme than the RDF values of mashes containing the same amount of nonmutated enzyme. B. Other Significant Accomplishment(s), if any. Determined additional gene sequences responsible for stability of barley alpha-glucosidase. C. Significant Accomplishments/Activities that Support Special Target Populations: none. 5.Describe your major accomplishments over the life of the project, including their predicted or actual impact? This project has resulted in the discovery of alpha-glucosidase's contributions to the in vitro hydrolysis of seed starch under optimal temperature and pH and has determined its contributions to starch hydrolysis under the more unfavorable conditions used by many industries. This project has led others to discover the importance of this enzyme in starch hydrolysis during seed germination and to collaborative studies that established the presence of a unique form of this enzyme in leaves. Studies of enzymes that degrade fructans have led to additional collaborative work that has demonstrated the roles of apoplastic fructans in the development of a cell's ability to withstand freezing. 6.What do you expect to accomplish, year by year, over the next 3 years? 2003: Complete studies of resistant starch and expand if appropriate. Complete development of thermostable alpha-glucosidase construct for insertion in malting barley. 2004: Complete characterization of second gene that encodes a carbohydrase crucial to the production of fermentable sugars. Initiate collaborative effort for development of transgenic malting barley. 2005: Establish the importance in starch degradation of a recently discovered carbohydrase. 7.What technologies have been transferred and to whom? When is the technology likely to become available to the end user (industry, farmer other scientist)? What are the constraints, if known, to the adoption durability of the technology? A patent is pending on the alpha-glucosidase gene sequences we identified as important in thermostability. Review Publications Muslin, E.H., Clark, S.E., Henson, C.A. The effect of proline insertions on the thermostability of a barley alpha-glucosidase. Protein Engineering. 2002. v. 15. p. 29-33. Clark, S.E., Hayes, P.M., Henson, C.A. Characterization of a barley ubiquitous beta-amylase. Plant Physiology Supplement. 2002. Abstract No. 755.