title: Expression, refolding, and autocatalytic proteolytic processing of the interleukin-1 beta-converting enzyme precursor.
abstract: The interleukin-1 beta-converting enzyme is a heterodimeric cysteine protease that is produced as a 45-kDa precursor. The full-length precursor form of the enzyme was expressed in Escherichia coli as insoluble inclusion bodies. Following solubilization and refolding of the 45-kDa protein, autoproteolytic conversion to a heterodimeric form containing 10- and 20-kDa subunits was observed. This enzyme had catalytic activity against both natural (interleukin-1 beta precursor) and synthetic peptide substrates. The inclusion of a specific inhibitor (SDZ 223-941) of the converting enzyme in the refolding mixture prevented proteolytic processing to the 10-/20-kDa form. Similarly, refolding under nonreducing conditions also prevented processing. Time course experiments showed that the 10-kDa subunit was released from the 45-kDa precursor before the 20-kDa subunit, implying that the N-terminal portion of the precursor is released last and may play a regulatory role.
authors.affiliation: Department of Biotechnology, Sandoz Pharma Ltd., Basel, Switzerland.
authors.forename: P
authors.initials: P
authors.lastname: Ramage
authors.forename: D
authors.initials: D
authors.lastname: Cheneval
authors.forename: M
authors.initials: M
authors.lastname: Chvei
authors.forename: P
authors.initials: P
authors.lastname: Graff
authors.forename: R
authors.initials: R
authors.lastname: Hemmig
authors.forename: R
authors.initials: R
authors.lastname: Heng
authors.forename: H P
authors.initials: HP
authors.lastname: Kocher
authors.forename: A
authors.initials: A
authors.lastname: Mackenzie
authors.forename: K
authors.initials: K
authors.lastname: Memmert
authors.forename: L
authors.initials: L
authors.lastname: Revesz
mesh_terms: D000595:Amino Acid Sequence; D020170:Caspase 1; D003546:Cysteine Endopeptidases / Q000737:chemistry / Q000378:metabolism*; D004792:Enzyme Precursors / Q000737:chemistry / Q000378:metabolism*; D008969:Molecular Sequence Data; D017510:Protein Folding
chemical_list: D004792:Enzyme Precursors; D003546:Cysteine Endopeptidases; D020170:Caspase 1