Identification of substrates for transglutaminase in Physarum polycephalum, an acellular slime mold, upon cellular mechanical damage.
Transglutaminases are Ca(2+)-dependent enzymes that post-translationally modify proteins by crosslinking or polyamination at specific polypeptide-bound glutamine residues. Physarum polycephalum, an acellular slime mold, is the evolutionarily lowest organism expressing a transglutimase whose primary structure is similar to that of mammalian transglutimases. We observed transglutimase reaction products at injured sites in Physarum macroplasmodia upon mechanical damage. With use of a biotin-labeled primary amine, three major proteins constituting possible transglutimase substrates were affinity-purified from the damaged slime mold. The purified proteins were Physarum actin, a 40 kDa Ca(2+)-binding protein with four EF-hand motifs (CBP40), and a novel 33 kDa protein highly homologous to the eukaryotic adenine nucleotide translocator, which is expressed in mitochondria. Immunochemical analysis of extracts from the damaged macroplasmodia indicated that CBP40 is partly dimerized, whereas the other proteins migrated as monomers on SDS/PAGE. Of the three proteins, CBP40 accumulated most significantly around injured areas, as observed by immunofluoresence. These results suggested that transglutimase reactions function in the response to mechanical injury.
Wada
Fumitaka
F
Department of Applied Molecular Biosciences, Graduate School of Bioagricultural Sciences, Nagoya University, Chikusa, Nagoya 464-8601, Japan.
Hasegawa
Hiroki
H
Nakamura
Akio
A
Sugimura
Yoshiaki
Y
Kawai
Yoshiki
Y
Sasaki
Narie
N
Shibata
Hideki
H
Maki
Masatoshi
M
Hitomi
Kiyotaka
K
D000199:Actins / Q000302:isolation & purification / Q000378:metabolism; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001710:Biotin / Q000031:analogs & derivatives / Q000378:metabolism; D002103:Cadaverine / Q000031:analogs & derivatives / Q000378:metabolism; D002135:Calcium-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism; D000226:Mitochondrial ADP, ATP Translocases / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D016839:Physarum polycephalum / Q000378:metabolism*; D015800:Protozoan Proteins / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D013314:Stress, Mechanical; D011503:Transglutaminases / Q000378:metabolism*
D000199:Actins; C406924:CBP40 protein, Physarum; D002135:Calcium-Binding Proteins; D015800:Protozoan Proteins; D001710:Biotin; D000226:Mitochondrial ADP, ATP Translocases; D011503:Transglutaminases; D002103:Cadaverine