A Tale Of Two Subunits: Co-Expression Using The Dual Plasmid Approach Of The Two Subunit Protein Dimethylsulfide Monooxygenase From Hyphomicrobium Sulfonivorans
- ASU Author/Contributor (non-ASU co-authors, if there are any, appear on document)
- Lydia Harris (Creator)
- Institution
- Appalachian State University (ASU )
- Web Site: https://library.appstate.edu/
- Advisor
- Megen Culpepper
Abstract: Dimethylsulfide (DMS) is a volatile organic sulfur compound (VOSC) that makes up about 70% of the atmospheric sulfur flux. Its ability to increase cloud formation, thereby lowering solar radiation, makes it an important gas to investigate for climate cooling options. Dimethylsulfide monooxygenase is an enzyme that was purified and characterized from the soil bacteria Hyphomicrobium sulfonivorans (H. sulfonivorans) as a flavin mononucleotide dependent two-component monooxygenase for DMS degradation. The A subunit, DmoA, is a putative monooxygenase, and the B subunit, DmoB, is a putative flavin reductase. The sequence identity of the dmoA gene is known. However, there are two dmoB gene candidates on the same operon that encode for putative flavin reductases. Protein isolation and purification from H. sulfonivorans indicates that the DmoA and DmoB subunits coexpress. DmoA and one of the flavin reductase candidates, DmoB176, have been co-expressed using the two plasmid approach in BL21DE3 E. coli. The dual plasmids were co-transformed in BL21DE3 E. coli, expressed, and purified via affinity chromatography. Initial data confirms expression of the DmoA monooxygenase protein at Mr of 53 kDa and DmoB176 at Mr of 19 kDa. Current research is being performed to standardize the co-expression procedure in order to measure enzymatic activity.
A Tale Of Two Subunits: Co-Expression Using The Dual Plasmid Approach Of The Two Subunit Protein Dimethylsulfide Monooxygenase From Hyphomicrobium Sulfonivorans
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Created on 7/3/2018
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Additional Information
- Publication
- Honors Project
- Harris, L. (2018). "A Tale Of Two Subunits: Co-Expression Using The Dual Plasmid Approach Of The Two Subunit Protein Dimethylsulfide Monooxygenase From Hyphomicrobium Sulfonivorans." Unpublished Honors Thesis. Appalachian State University, Boone, NC.
- Language: English
- Date: 2018
- Keywords
- Dimethylsulfide monooxygenase,
Protein co-expression,
Molecular cloning,
CLAW Hypothesis,
Hyphomicrobium sulfonivorans