Expression, Purification and Characterization of Recombinant 2-(2'-hydroxyphenyl)benzenesulfinase desulfinase from Rhodococcus erythropolis sp. IGTS8
| dc.contributor.advisor | Watkins, Linette M. | |
| dc.contributor.author | Harper, Leanne T. | |
| dc.contributor.committeeMember | Lewis, L. Kevin | |
| dc.contributor.committeeMember | David, Wendi M. | |
| dc.date.accessioned | 2012-08-15T17:16:51Z | |
| dc.date.available | 2012-08-15T17:16:51Z | |
| dc.date.issued | 2011-08 | |
| dc.description.abstract | Catalytic biodesulfurization uses microbes to remove the heterocyclic sulfur from dibenzothiophene-(DBT) without degrading the hydrocarbon fuel value. Rhodococcus erythropolis has been determined to degrade DBT to the final products 2-hydroxybiphenyl and sulfite using four enzymes. The last metabolic step in the pathway is performed by 2-(2’-hydroxyphenyl)benzenesulfinate desulfinase(DszB) and is the rate-limiting step. The gene coding for the DszB from Rhodococcus erthropolis IGTS8 was cloned into the expression vector pTAC-MAT-Tag-2 and then transformed into BL21(DE3) containing pREP4-GroESL. The enzyme was successfully expressed and purified using the ProBondTM Nickel column. Quantification showed a yield of 0.10 mg of DszB from 500 g of wet cells. Fluorimetric studies showed optimal activity at 35 ºC and pH range 8-10. The temperature stability range was 25- 35 º C. Kinetic studies show a Km of 3.75 ± 4.79µM and kcat 0.68 ± 0.15 min-1, respectively. The recombinant HPBS desulfinase showed similar optima temperature and stability as the native enzyme. The kinetic parameters shared similar values with the native enzyme, within experimental error. | |
| dc.description.department | Chemistry and Biochemistry | |
| dc.format | Text | |
| dc.format.extent | 50 pages | |
| dc.format.medium | 1 file (.pdf) | |
| dc.identifier.citation | Harper, L. T. (2011). <i>Expression, purification and characterization of recombinant 2-(2'-hydroxyphenyl)benzenesulfinase desulfinase from Rhodococcus erythropolis sp. IGTS8</i> (Unpublished thesis). Texas State University-San Marcos, San Marcos, Texas. | |
| dc.identifier.uri | https://hdl.handle.net/10877/4327 | |
| dc.language.iso | en | |
| dc.subject | Desulfurization | |
| dc.subject | Rhodococcus erythropolis IGTS8 | |
| dc.subject | Recombinant expression system | |
| dc.subject | GroESL | |
| dc.subject | Chaperonine proteins | |
| dc.title | Expression, Purification and Characterization of Recombinant 2-(2'-hydroxyphenyl)benzenesulfinase desulfinase from Rhodococcus erythropolis sp. IGTS8 | |
| dc.type | Thesis | |
| thesis.degree.department | Chemistry and Biochemistry | |
| thesis.degree.discipline | Biochemistry | |
| thesis.degree.grantor | Texas State University | |
| thesis.degree.level | Masters | |
| thesis.degree.name | Master of Science |