Fusion proteins containing the starch-binding domains of glucoamylase and cyclodextrin glucanotransferase
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The function of the Department of Chemical and Biological Engineering has been to prepare students for the study and application of chemistry in industry. This focus has included preparation for employment in various industries as well as the development, design, and operation of equipment and processes within industry.Through the CBE Department, Iowa State University is nationally recognized for its initiatives in bioinformatics, biomaterials, bioproducts, metabolic/tissue engineering, multiphase computational fluid dynamics, advanced polymeric materials and nanostructured materials.
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The Department of Chemical Engineering was founded in 1913 under the Department of Physics and Illuminating Engineering. From 1915 to 1931 it was jointly administered by the Divisions of Industrial Science and Engineering, and from 1931 onward it has been under the Division/College of Engineering. In 1928 it merged with Mining Engineering, and from 1973–1979 it merged with Nuclear Engineering. It became Chemical and Biological Engineering in 2005.
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1913 - present
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- Department of Chemical Engineering (1913–1928)
- Department of Chemical and Mining Engineering (1928–1957)
- Department of Chemical Engineering (1957–1973, 1979–2005)
- Department of Chemical and Biological Engineering (2005–present)
- College of Engineering(parent college)
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Abstract
This work addresses the interactions of a starch-binding domain (SBD) from glucoamylase-I (GA-I) of Aspergillus niger with raw starch and a possible application of the SBD as an affinity tail for a single-step recovery and purification of fusion proteins from crude cell extracts, using native starch granules as an adsorbent;Our previous studies indicated that GA-I adsorbed specifically and strongly to raw corn starch primarily via its SBD, which is located at the C-terminus of the enzyme molecule. A [beta]-galactosidase ([beta]-gal) fusion protein (BSB119) containing the C-terminal 119 aminoacids from GA-I showed significantly higher affinity for starch than a [beta]-gal control. This fusion protein was used as a model system to study the adsorption and elution behavior of SBD from starch. The results showed that the SBD is an independent domain that is functional when attached to another protein and the fusion expressed in E. coli. The adsorbed fusion protein was purified by competitive elution with various oligosaccharides;To test the generality of the proposed fusion protein approach for protein purification, another fusion protein (GSB119) was constructed with the same 119 aminoacids of SBD and glutathione S-transferase. This fusion protein formed inclusion bodies when produced in E. coli. Several procedures to reduce inclusion body formation were examined but were not very successful. The fusion protein was tested for its starch-binding activity and found to bind specifically to starch and easily eluted with 10 mM [beta]-cyclodextrin;Cyclodextrin glucanotransferase (CGTase) from Bacillus macerans also contains a starch-binding domain called domain E which is about 60% homologous to the SBD from GA-I. A [beta]-gal fusion protein with domain E (BDE109) from CGTase was also constructed and produced in E. coli. The starch-binding activity of BDE109 was compared to that of BSB119. It was found that both the SBD and domain E have similar affinities for the starch granule but different saturation capacities.