The biochemical and physiological function of mannan and CELLULOSE - PowerPoint PPT Presentation

The biochemical and physiological function of mannan and CELLULOSE SYNTHASE LIKE D2 protein in Triticum endosperm Yves Verhertbruggen October 2014 Agreenskills Annual Meeting - Toulouse

Who? Where? How? What? What? What?

Context Albumen 83% 65-70 % feruloylated arabinoxylan (AX) 20-25 % (1-3)(1-4)- β -D Glucan (MLG) Outer layers 14% ??? ca. 7 % Mannan Embryo 2-4 % Cellulose 3% Trace of pectin (HG, RGI, RGII) Trace of Xyloglucan (XG)

Aims of the project Defining: 1. The structure Which one(s) ??? 2. The biological function 3. The biosynthesis of mannan in wheat endosperm

Aims of the project Defining: Why ??? 1. The structure Mannan occurrence 350°J 150°J 2. The biological function aleurone 3. The biosynthesis of mannan Albumen amylacé in wheat endosperm

Aims of the project Defining: 1. The structure GDP-D-mannose GDP mannan β - (1→4) - mannosyltransferase 2. The biological function How ??? Man-(Man) n -Man Man-Man-(Man) n -Man 3. The biosynthesis of mannan in wheat endosperm

Methodology 1. Biochemistry Enzymatic fingerprint HPLC/GC MS Xylanase (eau) Xylanase (eau) betaglucanase (eau) betaglucanase (eau) Mannan extract treated with a mannanase 500000 500000 Mannobiose G3 G3 G3 Mono mannose Absorbance (nC) Mannotriose DP5 DP5 DP5 DP5 400000 400000 X2 X2 X2 NaOAc 1M: 0.3% Flow: 0.4 ml/min 300000 300000 G4 G4 G4 X1 X1 X1 200000 200000 Time of retention (min) Wheat sample treated with a mannanase DP6 DP6 DP6 DP6 Absorbance (nC) 100000 100000 DP4 DP4 DP4 DP4 NaOAc 1M: 0.3% DP7 DP7 DP7 DP7 Flow: 0.4 ml/min 0 0 DP9 DP9 DP9 DP9 -100000 -100000 Time of retention (min) 0 0 5 5 10 10 15 15 20 20 25 25 30 30 35 35 Tim e ( m in) Tim e ( m in)

Methodology 2. Molecular biology Ta CSLD2 RNAi transformant Collaboration with R. Mitchell (Rothamsted Research, United Kingdom)

Methodology 2. Molecular biology Test of enzymatic activity Endogenous acceptors 400 Transient expression in Nicotiana benthamiana Activity (% of control) 300 200 100 0 3. Preparation of microsomes and affinity purification of GT 4. Incubation with donors and acceptors and detection of potential products

Impact 1. Milling and food industry 2. Biofuel industry 3. « How to feed a growing global population? »

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