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United States Department of Agriculture

Agricultural Research Service

Title: Two-Dimensional Analysis of Proteinase Activity

Author
item OPPERT, BRENDA

Submitted to: Journal of Biochemical and Biophysical Methods
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: March 2, 2006
Publication Date: July 1, 2006
Repository URL: http://www.sciencedirect.com
Citation: Oppert, B.S. 2006. Two-dimensional analysis of proteinase activity. Journal of Biochemical and Biophysical Methods 67: 173-179.

Interpretive Summary: Indianmeal moths that are resistant to Bacillus thuringiensis toxins (Bt) have different digestive proteinases than Indianmeal moths that are sensitive to Bt. To study these differences, a new biochemical technique was developed. Proteinases are separated based on their electric charge, then separated by their molecular mass. These "two-dimensionally" separated proteinases are then detected using a specific enzyme substrate in a previously developed activity-blot assay. The method permits detection of individual proteinases in a mixture without prior purification, which can be further studied for their contribution to insect digestion as well as resistance to Bt toxins.

Technical Abstract: Identification and characterization of proteinase activities and related physiological differences in Bacillus thuringiensis-susceptible and -resistant colonies of Plodia interpunctella will be helpful to develop and/or improve control strategies for this pest insect. To resolve further the differences in proteinase activities in B. thuringiensis-susceptible and -resistant P. interpunctella colonies, a function-based activity profiling method was developed by combining the resolution of two-dimensional electrophoresis with proteolytic activity detection assays. The technique was used to identify differences in proteinase activity patterns from enzymes isolated from two different populations of P. interpunctella.

Last Modified: 9/10/2014
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