Dissecting processing and apoptotic activity of a cysteine protease by mutant analysis.
We have compared the behavior of wild-type mouse NEDD-2, a neural precursor cell-expressed, developmentally down-regulated cysteine protease gene, to various mutant forms of the gene in both apoptotic activity in neuronal cells and proteolytic cleavage in the Semliki Forest virus and rabbit reticulocyte protein expression systems. Our results confirm that NEDD-2 processing and apoptotic activity are linked phenomena. They identify aspartate residues as likely targets for autocatalytic cleavage. They establish that cleavage events only occur at specific sites. Finally, they pinpoint differential effects of individual mutations on the overall proteolytic cleavage patterns, raising interesting questions related to the mechanisms of subunit assembly.
Geneva Biomedical Research Institute, Glaxo Wellcome Research and Development S.A., Switzerland.
B
B
Allet
A
A
Hochmann
I
I
Martinou
A
A
Berger
M
M
Missotten
B
B
Antonsson
R
R
Sadoul
J C
JC
Martinou
L
L
Bernasconi
D000818:Animals; D017209:Apoptosis; D016466:CHO Cells; D053143:Caspase 2; D020169:Caspases; D002460:Cell Line; D002470:Cell Survival; D003001:Cloning, Molecular; D006224:Cricetinae; D003545:Cysteine; D003546:Cysteine Endopeptidases / Q000378:metabolism*; D015870:Gene Expression; D051379:Mice; D016297:Mutagenesis, Site-Directed; D009474:Neurons / Q000166:cytology / Q000502:physiology*; D017354:Point Mutation; D016133:Polymerase Chain Reaction; D014176:Protein Biosynthesis; D011506:Proteins / Q000378:metabolism*; D011817:Rabbits; D011994:Recombinant Proteins / Q000378:metabolism; D012156:Reticulocytes / Q000378:metabolism; D012672:Semliki forest virus; D013564:Sympathetic Nervous System / Q000166:cytology / Q000502:physiology; D014162:Transfection
D011506:Proteins; D011994:Recombinant Proteins; D053143:Caspase 2; D020169:Caspases; D003546:Cysteine Endopeptidases; D003545:Cysteine