Probing the Mg2+-dependent optical property changes of the functionally unknown Middle Rhodopsin from Haloquadratum walsbyi, HwMR

Autor: Ling-Ning Ko, 柯齡甯
Rok vydání: 2018
Druh dokumentu: 學位論文 ; thesis
Popis: 106
In haloarchaeas, microbial rhodopsins (M-Rho) are vital proteins for solar energy capture, as well as seeking for optimal living environment. Based on different physiological functions and mechanisms, two types of M-Rho have been found: light-driven ion transporters (bacteriorhodopsin, BR and halorhodopsin, HR) and phototaxis receptors (sensory rhodopsin, SR). Haloquadratum walsbyi (Hw) survives a 2.0 M MgCl2 condition and three M-Rho were identified in its genome, namely light-driven ion transporters HwBR, HwHR, and the functionally unknown HwMR. Previous studies showed HwMR to have the conserved residues from both BR and SRII; it even possesses photochemical properties of both, too. Based on our previous findings that HwBR was acid-tolerant and HwHR had extremely high affinity to chloride. Since Hw cells were found to survive an otherwise harmful 2.0 M Mg2+ environment, the goal of this study was set to seek the specific ion species, including Mg2+, that associate with MR. Here, we present the maximum absorbance (Abs-max) variations of three Hw M-Rho proteins under different ion species and concentrations and ion-dependent photocycle kinetics were recorded. Our results showed only MR appeared to be Mg2+ and Na+ sensitive among three M-Rhos in Hw. Furthermore, only the photocycle kinetics of MR showed significant changes under Mg2+. In order to investigate the critical site(s) related to Mg2+ binding, we mutated several amino acids in MR, and concluded that D84 which locates on the extracellular half of protein contributed to ion selectivity; the T216 near the retinal binding pocket, as well as cytoplasmic side D95 might be the specific Mg2+ binding sites. Further physiological significance is under study.
Databáze: Networked Digital Library of Theses & Dissertations
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