Aldehyde oxidase in wild type and aba1 mutant leaves of Nicotiana plumbaginifolia

Abstract

Aldehyde oxidase (AO; EC 1.2.3.1) activity was measured in rosette leaves of the wild type and aba1 mutant (I217) of Nicotiana plumbaginifolia. An activity band was detected in the extract of the wild type by staining after gel electrophoresis using cinnamaldehyde as a substrate, but not in that of I217. However, after treatment with Na2S and dithionite, an AO-activity band was detected in the extract of I217 at the same position as that of the wild type extract. These results indicated that I217 had AO apoprotein but the last step of molybdenum cofactor biosynthesis, from nitrate reductase form (dioxo form) to hydroxylase form (desulfo form), was blocked. Since aba1 is known to be impaired in ABA synthesis, we examined whether the leaf AO is an abscisic aldehyde (ABAId) oxidase. AO was purified from the leaves of wild type plants. After several steps of purification using cinnamaldehyde as a substrate which has a structure similar to ABAld, a partially purified enzyme preparation with a purification factor of about 160-fold was obtained. The apparent molecular mass of AO was estimated to be approximately 290 kDa by gel filtration. The enzyme had a relatively wide substrate specificity for aldehydes including ABAId. The possible involvement of Nicotiana AO in ABA biosynthesis is discussed.