Purification and characterization of the RfpA phytochrome of a thermophilic cyanobacterium

Perceiving light is a crucial step for all photosynthetic organisms. This perception of light is mediated by a group of photoreceptor proteins called phytochrome that exhibit high responsiveness to light and govern other biological processes like growth and development in plants, bacteria, fungi, al...

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Main Author: Ahmed, Syeda Josna
Other Authors: Matemaattis-luonnontieteellinen tiedekunta, Faculty of Sciences, Bio- ja ympäristötieteiden laitos, Department of Biological and Environmental Science, Jyväskylän yliopisto, University of Jyväskylä
Format: Master's thesis
Language:eng
Published: 2024
Subjects:
Online Access: https://jyx.jyu.fi/handle/123456789/96420
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author Ahmed, Syeda Josna
author2 Matemaattis-luonnontieteellinen tiedekunta Faculty of Sciences Bio- ja ympäristötieteiden laitos Department of Biological and Environmental Science Jyväskylän yliopisto University of Jyväskylä
author_facet Ahmed, Syeda Josna Matemaattis-luonnontieteellinen tiedekunta Faculty of Sciences Bio- ja ympäristötieteiden laitos Department of Biological and Environmental Science Jyväskylän yliopisto University of Jyväskylä Ahmed, Syeda Josna Matemaattis-luonnontieteellinen tiedekunta Faculty of Sciences Bio- ja ympäristötieteiden laitos Department of Biological and Environmental Science Jyväskylän yliopisto University of Jyväskylä
author_sort Ahmed, Syeda Josna
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description Perceiving light is a crucial step for all photosynthetic organisms. This perception of light is mediated by a group of photoreceptor proteins called phytochrome that exhibit high responsiveness to light and govern other biological processes like growth and development in plants, bacteria, fungi, algae, and cyanobacteria. They utilize red light and far-red light to regulate different light-dependent cellular processes. Certain cyanobacteria contain a distinct phytochrome called RfpA, which enables them to carry out photosynthesis by harnessing far-red light. So far there is no structural-functional relationship is known about this protein. The aim of this study was to purify RfpA to explore its signaling mechanism and spectral characterization to observe the photocycle of the phytochrome. Due to the difficulties in the purification of full-length protein, we constructed three different versions of the protein varying in length and containing different domains (GAF-PHY-PAS, GAF-PHY, and GAF only) using site-directed mutagenesis. Although our primary goal was the purification of the full-length protein, we attempted to obtain the longest possible fragment of the protein. We successfully managed to purify GAF-PHY and GAF mutants by affinity and size exclusion chromatography. Furthermore, UV-Vis spectroscopy was performed to study the photocycle of the protein. Spectroscopy study highlights that RfpA protein exists predominantly in Pfr (Far-red light absorbing form) as its resting state, denoting the characteristics of a bathy phytochrome. These findings provide remarkable insight into the unique spectral properties of RfpA, which can be beneficial in developing optogenetic tools and biotechnological applications.
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spellingShingle Ahmed, Syeda Josna Purification and characterization of the RfpA phytochrome of a thermophilic cyanobacterium RfpA phytochrome Ekologia ja evoluutiobiologia Ecology and evolutionary biology 4011 proteiinit syanobakteerit yhteyttäminen proteins cyanobacteria photosynthesis
title Purification and characterization of the RfpA phytochrome of a thermophilic cyanobacterium
title_full Purification and characterization of the RfpA phytochrome of a thermophilic cyanobacterium
title_fullStr Purification and characterization of the RfpA phytochrome of a thermophilic cyanobacterium Purification and characterization of the RfpA phytochrome of a thermophilic cyanobacterium
title_full_unstemmed Purification and characterization of the RfpA phytochrome of a thermophilic cyanobacterium Purification and characterization of the RfpA phytochrome of a thermophilic cyanobacterium
title_short Purification and characterization of the RfpA phytochrome of a thermophilic cyanobacterium
title_sort purification and characterization of the rfpa phytochrome of a thermophilic cyanobacterium
title_txtP Purification and characterization of the RfpA phytochrome of a thermophilic cyanobacterium
topic RfpA phytochrome Ekologia ja evoluutiobiologia Ecology and evolutionary biology 4011 proteiinit syanobakteerit yhteyttäminen proteins cyanobacteria photosynthesis
topic_facet 4011 Ecology and evolutionary biology Ekologia ja evoluutiobiologia RfpA phytochrome cyanobacteria photosynthesis proteiinit proteins syanobakteerit yhteyttäminen
url https://jyx.jyu.fi/handle/123456789/96420 http://www.urn.fi/URN:NBN:fi:jyu-202407155246
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