Studies have revealed in plant chloroplasts, mitochondria, cell walls, and cytoplasm the existence of transglutaminase (TGase) activities, similar to those known in animals and prokaryotes having mainly structural roles, but no protein has been associated to this type of activity in plants. A recent computational analysis has shown in Arabidopsis the presence of a gene, AtPng1p, which encodes a putative N-glycanase. AtPng1p contains the Cys-His-Asp triad present in the TGase catalytic domain. AtPng1p is a single gene expressed ubiquitously in the plant but at low levels in all light-assayed conditions. The recombinant AtPng1p protein could be immuno-detected using animal TGase antibodies. Furthermore, western-blot analysis using antibodies raised against the recombinant AtPng1p protein have lead to its detection in microsomal fraction. The purified protein links polyamines-spermine (Spm) . spermidine (Spd) . putrescine (Put)-and biotin-cadaverine to dimethylcasein in a calcium-dependent manner. Analyses of the g-glutamyl-derivatives revealed that the formation of covalent linkages between proteins and polyamines occurs via the transamidation of g-glutamyl residues of the substrate, confirming that the AtPng1p gene product acts as a TGase. The Ca
21-and GTP-dependent cross-linking activity of the AtPng1p protein can be visualized by the polymerization of bovine serum albumine, obtained, like the commercial TGase, at basic pH and in the presence of dithiotreitol. To our knowledge, this is the first reported plant protein, characterized at molecular level, showing TGase activity, as all its parameters analyzed so far agree with those typically exhibited by the animal TGases.Transglutaminases (TGases; E.C. 2.3.2.13) catalyze protein cross-linking by the interaction of an acyl acceptor glutamyl residue and an amine donor, a lysyl residue, of the same or another protein, or the formation of a link between a protein and a free primary amine, like a polyamine (PA). The terminal aminogroups of PAs conjugate one or two glutamyl residues giving rise either to mono-(g-glutamyl)-polyamines (mono-PAs) or bis-(g-glutamyl)-polyamines (bis-PAs; Folk, 1980). The posttranslational cross-linking of proteins is one of the essential physiological processes involved in the stabilization of tissue and cellular matrices. High molecular protein networks can be generated by TGase catalysis when cross-links are formed between Gln and either Lys residues or PAs (bis-PA derivatives) thus forming bridges between different proteins. At present, both in eukaryotes and prokaryotes several TGases have been described exhibiting a large number of functions (Aeschlimann et al., 1998;Griffin et al., 2002;Lorand and Graham, 2003). Eight distinct TGase isoenzymes have been identified in different animal tissues, and some of these have been purified and characterized at molecular level (Aeschlimann et al., 1998;Griffin et al., 2002;Lorand and Graham, 2003).TGases are classified in three structural families: (1) the papain-like TGases, (2) the protein di...
