Identification of a 23 kDa protein from maize photoaffinity-labelled with 5-azido-[7-3H]indol-3-ylacetic acid

Abstract

A 23 kDa protein (p23) was identified in microsomal extracts from maize coleoptiles by photoaffinity labelling with 5-azido-[7-3H]indol-3-ylacetic acid ([3H]N3IAA). Labelling of p23 was blocked by unlabelled IAA, N3IAA, indol-3-ylbutyric acid and indol-3-yl-lactate. In addition, labelling was efficiently decreased by tryptophan, as well as by the scavenger p-aminobenzoic acid. Labelling was, however, not affected by synthetic auxins such as 1-naphthylacetic acid or 2,4-dichlorophenoxyacetic acid. Competition data suggest that the label was probably bound via the indole ring, and hence labelling was not specific for auxins. The 23 kDa protein was solubilized from crude microsomes by extraction with Triton X-100 and purified to homogeneity by ion-exchange, size-exclusion and reversed-phase chromatography. After electroblotting, the amino acid sequences of the p23 N-terminus as well as the several tryptic peptides were obtained. Database comparisons revealed sequence identity with a maize manganese superoxide dismutase. We conclude that photoaffinity labelling of p23 was pseudo-affinity, and therefore the binding site for IAA is not specific.

Cloning of Genes Encoding Auxin-Binding Proteins (ABP19/20) from Peach: Significant Peptide Sequence Similarity with Germin-Like Proteins

“…ABP1 isolated from maize coleoptile is the best characterized ABP and is probably involved in mediating rapid auxin effects on cell elongation (Napier and Venis 1991). Some ABPs have been identified as proteins with enzymatic activities such as /?-glucosidase , Brzobohaty et al 1993), 1,3-0-glucanase (Macdonald et al 1991), glutathione S-transferase (Bilang et al 1993, Zettl et al 1994, manganese superoxide dismutase (Feldwisch et al 1995), and glutathione-dependent formaldehyde dehydrogenase Abbreviations: ABP, auxin-binding protein; ORF, open reading frame.…”

A new perspective on auxin perception

“…Early biochemical investigations identified a number of auxinbinding soluble proteins such as 1,3-glucanase (MacDonald et al, 1991), ß-glucosidase (Campos et al, 1992), glutathione S-transferase (Bilang et al, 1993) and superoxide dismutase (Feldwisch et al, 1994). Two soluble ABPs with a relatively low affinity for IAA were purified and reported to stimulate RNA synthesis in isolated nuclei (Kikuchi et al, 1989).…”