Crystal structure of FIP1 and its cooperative activity with FIN219/JAR1
Date Issued
2012
Date
2012
Author(s)
Chen, Chun-Yen
Abstract
Light signal plays an important role in plant growth and development. Photomorphogenesis and skotomorphogenesis are plant physiological mechanisms developed for adapting to light, including light fluence, photoperiodism, phototropism, shade avoidance and circadian rhythms. Jasmonate is a crucial phytohormone. It can be accumulated in the wounded or infected tissue and triggered defense response. Recent study confirms that jasmonate regulates root formation, flower development and fruit maturation, it affects deeply the reproductive physiology.
FIN219 (Far-red insensitive 219), also known as JAR1 (Jasmonate Resistant 1), involved in far-red light signaling pathway and possessed catalytic activity in conjugating jasmonic acid to isoleucine. In addition, FIP1 (FIN219-interacting protein 1) screened from yeast two-hybrid system also participated in far-red light signaling pathway. It belongs to the member of Arabidopsis thaliana AtGSTU family and possesses GST enzyme activity, previous study identified that FIN219 interacts with FIP1, then FIP1 improves FIN219 stability and regulates FIN219 enzyme activity.
This study aimed to resolve FIN219 and FIP1 protein structure, function and enzyme activity. FIN219 and FIP1 proteins were expressed in Escherichia coli system and further purified using gel filtration chromatography. Interestingly, FIN219 could interacts with SjGST, a GST protein expressed from vector pGEX 4T-1, and interfered the ability of SjGST binding to GST affinity column. The FIN219-SjGST complex could be observed in gel filtration chromatography. The binding affinities and enzymatic kinetics between FIN219 and FIP1, and between FIN219 and SjGST were determined by Quartz Crystal microbalance (QCM). The results of kinetics of FIP1 demonstrated that FIP1 would promote maxmium velocity (Vmax) and catalytic constant (Kcat) of FIN219. Finally we screened FIN219-GST, FIN219-FIP1 and FIP1 crystallization condition. Only the crystals of FIP1 were obtained. The resolved structure of FIP1 is a dimer and possesses two active sites, with highly conserved residues Glu and Ser in G-site. In addition, FIP1 contained a larger H-site in comparison to that of GmGSTU4-4. It exhibited the importance of GST tau class in detoxification and in metabolism of herbicide compounds.
Subjects
Jasmonate
Type
thesis
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