A COMPLEX ARRAY OF PROTEINS RELATED TO THE MULTIMERIC LEUCINE AMINOPEPTIDASE OF TOMATO

Leucine aminopeptidase (LAP) mRNAs are induced in response to mechanic al wounding, pathogen infection, and insect infestation (V. Pautot, F. M. Holzer, B. Reisch, L.L. Walling [1993] Proc Natl Acad Sci USA 90: 9 906-9910). Polyclonal antibodies to a glutathione S-transferase-LAP fu sion protein and affinity-purified antibodies recognizing LAP antigeni c determinants detected four classes of polypeptides in tomato (Lycope rsicon esculentum) leaves. All four classes had multiple polypeptides in two-dimensional polyacrylamide gel electrophoresis immunoblots. Alt hough antigenically related to the wound-induced tomato LAP proteins, the 77- and 66-kD LAP-like proteins accumulated in both healthy and wo unded leaves. Two classes of 55-kD polypeptides with distinctive isoel ectric points were designated as plant LAPs; only the acidic LAP prote ins accumulated to high levels after mechanical wounding or Pseudomona s syringae pv tomato infection of tomato leaves. The temporal accumula tion of LAP mRNAs was correlated with the increase in acidic LAP prote in subunits. A slow-migrating LAP activity was detected using a native gel assay after wounding. The molecular mass of the native wound-indu ced LAP enzyme was 353 kD. The 55-kD acidic LAP proteins were associat ed with induced LAP activity, whereas the neutral LAPs and the LAP-lik e proteins were not associated with this exopeptidase. A second, fast- migrating aminopeptidase was detected in both healthy and wounded toma to leaves. Cell fractionation experiments revealed that wound-induced LAP is a soluble enzyme.