dc.contributor.advisor | Essenberg, Richard C. | |
dc.contributor.author | Gupta, Snehalata | |
dc.date.accessioned | 2014-04-15T18:38:08Z | |
dc.date.available | 2014-04-15T18:38:08Z | |
dc.date.issued | 2004-07-01 | |
dc.identifier.uri | https://hdl.handle.net/11244/8540 | |
dc.description.abstract | Insect phenoloxidase (PO) plays an important role in melanization, wound healing and cuticle sclerotization. It is generated from prophenoloxidase (proPO) by proPO-activating proteinase (PAP) and two serine proteinase homologs (SPHs) at the end of a proteinase cascade. While partial isolation of PAP-1 from the cuticles of Manduca sexta prepupae was reported previously, we have purified PAP-1 to near homogeneity by hydroxyapatite, dextran sulfate, gel filtration, and lectin affinity chromatography. The KM and Vmax of PAP-1 towards IEARpNA did not change after the SPHs were added. The KM of PAP-1 towards proPO (in the presence of SPHs) was determined to be 16.5 6.0g/ml, close to proPO concentration in the larval hemolymph. PAP-1 itself cleaved more than 50% of proPO at Arg51 without generating any significant amount of PO activity. In the presence of SPHs, the cleavage of proPO by PAP-1 was moderately enhanced whereas the PO activity increase was more than 25 fold, indicating that the cleavage of proPO does not directly correlate with PO activity. However, proteolytic cleavage is critical for proPO activation since SPHs and APMSF-treated PAP-1 did not generate active PO by interacting with proPO. After over 20% of proPO was cleaved by PAP-1 which was then inactivated by M. sexta serpin-1J or APMSF, incubating the reaction mixture with SPHs failed to generate active PO either. In other words, SPHs can not generate PO activity by simply binding to cleaved proPO. Rather, proPO activation in M. sexta requires active PAP-1 and SPHs at the same time one for limited proteolysis and the other perhaps as a cofactor. | |
dc.format | application/pdf | |
dc.language | en_US | |
dc.publisher | Oklahoma State University | |
dc.rights | Copyright is held by the author who has granted the Oklahoma State University Library the non-exclusive right to share this material in its institutional repository. Contact Digital Library Services at lib-dls@okstate.edu or 405-744-9161 for the permission policy on the use, reproduction or distribution of this material. | |
dc.title | Thermobarometric Constraints on the Evolution of the Menderes Massif (Western Turkey): Insights into the Metamorphic History of a Complexly-Deformed Region | |
dc.type | text | |
dc.contributor.committeeMember | Jiang, Haobo | |
dc.contributor.committeeMember | Matts, Robert | |
osu.filename | Gupta_okstate_0664M_1040.pdf | |
osu.college | Agricultural Sciences and Natural Resources | |
osu.accesstype | Open Access | |
dc.description.department | Department of Biochemistry and Molecular Biology | |
dc.type.genre | Thesis | |