9 March 2016 The application of low angle light scattering to evaluate qualitatively and quantitatively the dynamics of formation of oligomers in heme protein sensors
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Abstract
The aim of this study is to investigate the structural organization and oligomerization properties of the sensory kinase protein DevS using low-angle light scattering (LALS) and gel filtration chromatography (HPLC). In addition, the structural characteristics of FixL and BSA were investigated and compared with DevS to better elucidate LALS technique. DevS is a direct and specific O2 sensing protein in Mycobacterium tuberculosis and acts as an activator of the transcription factor protein DevR. This latter triggers the latency state of tuberculosis under hypoxic conditions. DevS has been briefly evaluated under different conditions of concentration, ionic strength and temperature. LALS and gel filtration (HPLC) analysis were performed right after DevS purification process. The results of LALS for BSA proved to be highly reliable with a Rh value of c.a. 3.7 nm. Considering BSA a globular protein, the molecular weight estimative, using LALS was near 67 KDa, which is reasonably within the value reported in the literature. Preliminary LALS results showed a hydrodynamic radius (Rh) varying from 4.2-15.0 nm for DevS protein, and an average of 6.7 nm. These data supported, along with gel filtration, a dimer (~130 KDa) and tetramer (255 KDa) as the main DevS species. Additionally, it was found higher oligomeric species by gel filtration suggesting either an equilibrium of oligomers or an aggregation process that deserves further studies.
Conference Presentation
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Luis G. Sabino, Luis G. Sabino, Wellinson Gadelha Guimarães, Wellinson Gadelha Guimarães, Pedro Mikael Costa, Pedro Mikael Costa, Marta S. P. Carepo, Marta S. P. Carepo, Ana C. S. Gondim, Ana C. S. Gondim, Luiz G. F. Lopes, Luiz G. F. Lopes, Eduardo H. S. Sousa, Eduardo H. S. Sousa, "The application of low angle light scattering to evaluate qualitatively and quantitatively the dynamics of formation of oligomers in heme protein sensors", Proc. SPIE 9719, Biophysics, Biology, and Biophotonics: the Crossroads, 97190B (9 March 2016); doi: 10.1117/12.2213733; https://doi.org/10.1117/12.2213733
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