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Characterization of TaXPol-1, a Xylan Synthase Complex from Wheat
Jiang, Nan

2015, Doctor of Philosophy (PhD), Ohio University, Environmental and Plant Biology (Arts and Sciences).
It is believed that Golgi-localized, multi-protein complexes synthesize plant hemicellulosic polysaccharides. However, such complexes are not well characterized in plants, which is in sharp contrast with mammalian and yeast cells. This dissertation describes the characterization of the first hemicellulose synthase complex involved in xylan biosynthesis in wheat (named a Xylan Polymerase-1, TaXPol-1).

Xylan is the most abundant plant hemicellulosic polysaccharide on Earth and plays an important role in the integrity of cell walls. It consists of a ß-(1,4)-linked xylose backbone that can be substituted with a-(1,2)-D-glucuronic acid residues and/or a-L-arabinofuranosyl residues at the C-2 and/or C-3 positions of xylose residues.

A xylan synthase activity was immuno-purified from etiolated wheat seedlings and its protein composition determined by proteomics analysis. Four glycosyltransferases (GTs) were identified: CAZy GT43 family member (named TaGT43-4), GT47 family member (named TaGT47-13), and two GT75 family members (named TaGT75-3 and TaGT75-4). In addition, two new proteins were identified: a germin-like protein (TaGLP) belonging to the cupin family and a 26kDa protein annotated as vernalization-related 2 (TaVER2).

Heterologous co-expression of TaGT43-4, TaGT47-13, TaGT75-3, TaGT75-4, and TaGLP in Pichia pastoris cells resulted in reconstitution of xylan synthase activity in micosomes from transgenic yeast lines. The results showed that (i) TaGT43-4 and TaGT47-13 form a central core that is necessary and sufficient for synergistic incorporation of xylose and glucuronic acid, but requires TaGT75-3 and/or TaGT75-4 (putative mutases) for synergistic incorporation of xylose and arabinose, and (ii) TaGLP was needed for optimal xylan synthase activity. Furthermore, radio labeled products generated by microsomes from transgenic yeast cells have xylan characteristics.

Bimolecular fluorescence complementation (BiFC) and confocal microscopy were used to show that TaGT43-4 interacts with TaGT47-13, TaGT75-3, TaGT75-4, and TaGLP, and these proteins assemble into a complex in the endoplasmic reticulum (ER) before export to the trans-Golgi. Importantly, ER export of TaXPol-1 requires that TaGT43-4 and TaGT47-13 interact.
Faik Ahmed (Advisor)
Showalter Allan (Committee Member)
Kieliszewski Marcia (Committee Member)
Held Michael (Committee Member)
205 p.

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Jiang, N. (2015). Characterization of TaXPol-1, a Xylan Synthase Complex from Wheat. (Electronic Thesis or Dissertation). Retrieved from https://etd.ohiolink.edu/

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Jiang, Nan. "Characterization of TaXPol-1, a Xylan Synthase Complex from Wheat." Electronic Thesis or Dissertation. Ohio University, 2015. OhioLINK Electronic Theses and Dissertations Center. 22 Nov 2017.

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Jiang, Nan "Characterization of TaXPol-1, a Xylan Synthase Complex from Wheat." Electronic Thesis or Dissertation. Ohio University, 2015. https://etd.ohiolink.edu/

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