Gamma-glutamyltranspeptidase of Helicobacter pylori induces mitochondria-mediated apoptosis in AGS cells.
Gamma-glutamyltranspeptidase (GGT) is a novel protein involved in the induction of Helicobacter pylori-mediated apoptosis; however, the signal pathway involved in GGT-induced apoptosis remains unclear. Using DNA recombination techniques, ggt was cloned into pET117b and transformed into Escherichia coli. Recombinant GGT was purified using nickel-affinity resin and was digested by thrombin. Recombinant GGT induced apoptosis in AGS cells in a time-dependent manner, which was confirmed by TUNEL staining, the MTT assay and immunoblot analysis for caspases-9, -3, Bax, Bcl-2, Bcl-xL and cytochrome c release. Activation of caspase-3 and -9 following exposure to GGT increased in a time-dependent manner and upregulation of proapoptotic Bax and a downregulation of antiapoptotic Bcl-2 and Bcl-xL was detected. Apoptotic signals also trigger changes in mitochondria, which lead to a release of cytochrome c into the cytosolic space. The GGT-deficient mutant was not as able to induce apoptosis as the wild-type strain. These results indicate that GGT of H. pylori induces apoptosis via a mitochondria-mediated pathway.
Kim
Kyung-Mi
KM
Department of Microbiology, Gyeongsang National University College of Medicine, Chiram-dong 90, Jinju, Gyeongsangnam-do 660-751, Republic of Korea.
Lee
Seung-Gyu
SG
Park
Min-Gyu
MG
Song
Jea-Young
JY
Kang
Hyung-Lyun
HL
Lee
Woo-Kon
WK
Cho
Myung-Je
MJ
Rhee
Kwang-Ho
KH
Youn
Hee-Shang
HS
Baik
Seung-Chul
SC
D000230:Adenocarcinoma / Q000473:pathology*; D017209:Apoptosis / Q000502:physiology*; D001483:Base Sequence; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D016480:Helicobacter pylori / Q000201:enzymology*; D006801:Humans; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000502:physiology*; D011994:Recombinant Proteins / Q000302:isolation & purification; D013274:Stomach Neoplasms / Q000473:pathology*; D014407:Tumor Cells, Cultured; D005723:gamma-Glutamyltransferase / Q000302:isolation & purification / Q000378:metabolism*
D017931:DNA Primers; D011994:Recombinant Proteins; D005723:gamma-Glutamyltransferase