Chaperone-like properties of the prodomain of TNFalpha-converting enzyme (TACE) and the functional role of its cysteine switch.
The prodomain of TACE [TNFalpha (tumour necrosis factor alpha)-converting enzyme] is essential for the secretion of the functional enzyme. Previously, we showed that a TACE truncate was not secreted in the absence of the prodomain and that it was subjected to intracellular degradation. In the present study, we show that full-length TACE was also degraded when expressed without the prodomain. We demonstrate that the prodomain can rescue TACE's secretion in trans, suggesting an intramolecular chaperone function. We addressed the question whether a cysteine switch consensus motif is needed for the secretion of active TACE. The cysteine switch mutants [C184A (Cys184-->Ala)] of TACE resembled the wild-type functionally and in their sensitivity to inhibitors. Interestingly, TACE zymogen forms expressed in the context of the C184A mutation were susceptible to intracellular degradation, suggesting that the prodomain-bound TACE zymogen may be more accessible to intracellular proteinases when compared with mature TACE. Two independent findings confirmed that the catalytic domain of TACE is in a more open state when bound to its prodomain: (i) core tryptophan residues were exposed to the solvent in the procatalytic domain complex and (ii) LysC rapidly proteolysed the procatalytic domain complex but not mature TACE. Therefore the prodomain of TACE is a specific intramolecular chaperone that aids in the secretion of this enzyme, while keeping the catalytic domain in a relatively open conformation. The cysteine switch of TACE is not essential for the secretion of the functional enzyme, but may prevent intracellular degradation of the TACE zymogen.
Leonard
Jennifer D
JD
Department of Biochemistry and Biophysics and Johnson Research Foundation, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA.
Lin
Frank
F
Milla
Marcos E
ME
D051722:ADAM Proteins / Q000096:biosynthesis* / Q000737:chemistry* / Q000235:genetics; D000072198:ADAM17 Protein; D000595:Amino Acid Sequence; D003545:Cysteine / Q000737:chemistry; D004789:Enzyme Activation; D005786:Gene Expression Regulation / Q000502:physiology*; D018832:Molecular Chaperones / Q000737:chemistry*; D009154:Mutation; D017434:Protein Structure, Tertiary
D018832:Molecular Chaperones; D051722:ADAM Proteins; D000072198:ADAM17 Protein; D003545:Cysteine