IS PLANT TRANSGLUTAMINASE A BIFUNCTIONAL ENZYME?

A gene having the typical catalytic triad (His-Cys-Asp) of animal and bacteria transglutaminase (TGase) family, but without a significant sequence homology, has been identified in Arabidopsis thaliana genome. The recombinant first plant protein expressed by this gene, (with orthologs in rice, pear and apple), even though classified as a peptide:N-glycanase (AtPng1p), showed transglutaminase activity (Della Mea et al., 2004), presenting the main biochemical characteristics (pH, calcium- and GTP-dependence, substrate recognition, production of glutamyl-polyamine derivatives etc.) of animal TGases. It is expressed at very low levels during the normal development of the plant. There are no evidences of the supposed peptide:N-glycanase activity in Arabidopsis. The yeast orthologue of AtPng1 (ScPng1) is a bona fide PNGase (Suzuki, et al., 2006) but does not exhibit TGase activity. We replaced the catalytic domain of PNGase with the catalytic domain of AtPng1 to see if we can observe PNGase activity, but no PNGase activity can be detected, despite the high homology of TGase domain. Through systematic mutagenesis study, we identified two amino acid substitutions in AtPng1 TGase domain, which render the Sc protein inactive for PNGase activity. We also did not observe any complementation of in vivo PNGase activity when AtPng1 was expressed in yeast. These results suggest that Png1 may evolve into proteins with distinct functions at least between yeast and plants. An AtPng1p knock-out Arabidopsis strain presents different morphological and biochemical characteristics, namely a juvenile phenotype, smaller dimensions of the stem cells, when compared to those of the wild type. To magnify the differences in TGase activity, both strains have been subjected to thermal or wound stresses: Wt responds to wounding in short time, whereas KO is much less responsive. The subcellular localisation of the enzyme showed that more than one form should be present: a strong activity was detected in Wt- and KO-strain chloroplasts, and a weak or null activity respectively in microsomes. In the cell wall the presence of a TGase activity, possibly related to reparation of the wound, is suggested. Della Mea et al. (2004) Plant Physiol., 135: 2046-2054. Suzuki et al. (2006) J. Biol. Chem. 281, 22152-22160. Funds: PRIN 2007 and Progetto Strategico d’Ateneo 2006 to D.S-F. Grant for M. Della Mea supported by COE (Center-of-Excellence) Program of Osaka University.