Analysis of the structure and function of a cellulose binding domain

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Project objective

Research objectives and content Recent studies have demonstrated that cellulose binding domains (CBDs) play an important role in the activity of plant cell wall hydrolase against insoluble substrates. Pseudomonas enzymes consist of two distinct CBDs; a type II CBD domain and a second smaller domain. Although recent studies have shown that removal of both CBDs decreases the activity of Pseudomonas xylanase A (XYLA) against cellulose/hemicellulose complexes, it is unclear why Pseudomonas enzymes contain two distinct CBDs. The objective of this proposal is to determine the properties of the 40 residue CBD of XYLA and its role in the action of XYLA against the plant cell wall. Specifically, site-directed mutagenesis will be used to change the conserved aromatic residues to alanine to establish whether they play a pivotal role in binding cellulose, or are integral to the correct folding of the CBD. To establish the importance of the CBD in the activity of XYLA, xynA will be engineered such that it encodes two derivatives of XYLA in which either the large type II CBD (XYLA') or the small CBD (XYLA) has been removed. Analysis of the capacity of XYLA, XYLA' and XYLA" to hydrolyse cellulose/xylan complexes will establish why Pseudomonas glycosyl hydrolase, in general, have two distinct CBDs. Training content (objective, benefit and expected impact) This project will provide me with an excellent training in the use of recombinant DNA technology (site-directed mutagenesis, PCR, DNA sequencinge etc) to study the structure and function of proteins. I already have experience in the use of both biochemical and chemical techniques to study industrially important enzymes. At the completion of my training I will be able to use recombinant DNA technology to produce large quantities of these proteins, and engineer their structure to underpin more sophisticated studies on these important biocatalysts. Links with industry / industrial relevance (22) Professor Gilbert has strong industrial contacts with Zeneca BioProducts (enzymes in the Paper Pulp Industry), Thomas Swan (Paper pulp Industry), Genacor (Detergent Industry), Dalgety and FinnFeeds International (Animal Feeds and the Food industry). "

Original text from CORDIS.

Participants

• UNIVERSITY OF NEWCASTLE UPON TYNE · NEWCASTLE UPON TYNECoordinatorUnited Kingdom
• Not availableCity levelFrance

Links

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