IDENTIFICATION AND LOCALIZATION OF AN ACTIVE CUTINASE IN THE POLLEN OF BRASSICA-NAPUS L

Polyclonal antiserum and monoclonal antibodies raised to a purified cu tinase from Fusarium solani f. sp. pisi have been used to identify an active cutinase in the pollen of Brassica napus. These antibodies reco gnized a polypeptide with an estimated molecular weight of 22-kDa - a molecular weight indentical to that of the Fusarium cutinase - and loc alized this polypeptide to the intine of the pollen wall. Enzyme assay s on the renatured 22-kDa polypeptide after electroelution from a prep arative SDS-PAGE gel revealed the polypeptide to be an enzyme capable of catalysing the hydrolysis of tritiated apple cutin and the syntheti c substrate p-nitrophenyl butyrate. The molecular weight, immunologica l properties and substrate specificity of the Brassica cutinase sugges t that this enzyme resembles more closely fungal cutinases than it doe s the cutinase from the pollen of Nasturtium (Tropaeolum majus) - the only angiosperm cutinase so far characterized (Maiti et al., 1979, Arc h. Biochem. Biophys. 196, 412-423). These differences between the poll en cutinases from two members of the Dicotyledoneae are unexpected and predict a diversity of this class of pollen enzyme within the angiosp erms.