25-30 August 2019
Henry Ford Building
Europe/Berlin timezone

Using in cell 2D 13C-13C solid-state NMR as a biophysical tool to characterize cell-wall and starch from the microalga C. reinhardtii

Not scheduled
4h
Harnack House and Henry Ford Building

Harnack House and Henry Ford Building

Board: 374
Poster Posters

Speaker

Alexandre Poulhazan (Department of Chemistry, Université du Québec à Montréal)

Description

Saccharides, in addition to their important energetic role, are used in the cell as building blocks for complex molecules. Glycosidic bonds between them can create different types of molecules, with linear or complex branched assemblies.
The main storage molecule in the microalga C. reinhardtii is starch, one of the most abundant polysaccharide in nature. This glucose polymer is made of amorphous and crystalline domains which can be mainly found into two different structures: A and B-types. These structures have specific physicochemical properties and are valued differently in drug formulation, for example. Using high-resolution solid-state NMR Magic-Angle spinning (MAS) methods on 13C-labelled starches, we can assign starch and its constituents (amylopectin and amylose) in the two crystalline forms and in the amorphous state. 2D-INADEQUATE experiments enable the assignment of hereto unreported non-reducing end groups, and the assessment of starch chain length, crystallinity and hydration. Solid-state NMR can also inform us on dynamics and conformational disorder, in particular in the amorphous state.
C. reinhardtii also has an extracellular cell-wall mainly made of hydroxyproline-rich glycoproteins, with characteristics shared by both higher plants and mammalian extracellular matrix. Using additional 2D solid-state NMR methods on 13C labelled cell-wall extracts, we assign several amino acids, saccharides and links between them, revealing both molecular and dynamic complexity in the cell wall. Moreover, highly crystalline, amorphous and hydrated regions could be differentiated.
This work illustrates how high-resolution solid-state NMR can be used to enable the detection and identification of starch and cell-wall in situ in intact cells, therefore eliminating time consuming and potentially altering purification steps. Furthermore, we show how these NMR methods can be used to get detailed information on organic disordered solids, and even in living microorganisms, making in situ solid-state NMR a powerful tool to study molecules directly in the cell.

Primary authors

Alexandre Poulhazan (Department of Chemistry, Université du Québec à Montréal) Dr Alexandre A Arnold (Department of Chemistry, Université du Québec à Montréal) Prof. Dror E Warschawski (Laboratoire de Biologie Physico-Chimique des Protéines Membranaires, UMR 7099, CNRS, Université Paris Diderot and IBPC) Prof. Isabelle Marcotte (Department of Chemistry, Université du Québec à Montréal)

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