INFLUENCE OF EXPANSIN CARBOHYDRATE BINDING MODULE ON IN VITRO POLYGALACTURONASE ENZYME ACTIVITY

NARDI, C; ESCUDERO, C; VILLARREAL NM; MARTÍNEZ GA; CIVELLO PM

Tucumán

Congreso; XLV Reunión Anual SAIB; 2009

SAIB

Fruit softening is associated with cell wall disassembly mediated bythe action of a set of enzymes and proteins. EXPANSINs (EXPs), agroup of proteins with unknown enzymatic activity, andPOLYGALACTURONASEs (PGs) are proposed to be involved inthis process. It has been suggested that EXPs and PGs couldcooperatively participate in ripening-associated cell walldegradation, though there is scarce experimental evidence thatsupport such statement.FaEXP2 encodes a putative strawberry expansin (FaEXP2) with apredicted carbohydrate binding module (CBM) in its C-terminal andan “expansin domain” in its N-terminal.In order to evaluate the influence of expansin, we performed in vitroPG activity assays in the presence of full FaEXP2 or its CBM (CBMFaEXP2).Both cDNAs were cloned, over-expressed in E. coli, andthe recombinant polypeptides were purified. In vitro PG activity wasmeasured using enzymatic extract from tomato, citrus pectin assubstrate and the addition of FaEXP2 or full CBM-FaEXP2.The presence of CBM/FaEXP2 in the reaction mixture decreasedPG activity, and a slighter effect was found in the case of FaEXP2also. These results suggest that CB-FaEXP2 could interact withpectins and then interfere with the activity of polygalacturonaseenzyme