Purification of an interleukin-1?? converting enzyme-related cysteine protease that cleaves sterol regulatory element-binding proteins between the leucine zipper and transmembrane domains

  • Wang X
  • Pai J
  • Wiedenfeld E
 et al. 
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We describe the characterization and purification of a protease that cleaves sterol regulatory element-binding protein-1 {(SREBP-1)} and {SREBP-2} in vitro. Cleavage occurs between the basic helix-loop-helix-leucine zipper and the first transmembrane domain of each {SREBP.} This is the region in which the {SREBPs} are cleaved physiologically by a sterol-regulated protease that releases an {NH2-terminal} fragment that activates transcription of the genes for the low density lipoprotein receptor and 3-hydroxy-3-methylglutaryl {CoA} synthase. The cleavage enzyme, designated {SREBP} cleavage activity {(SCA),} belongs to a new class of cysteine proteases of the interleukin-1 beta-converting enzyme {(ICE)} family, all of which cleave at aspartic acid residues. Like {ICE,} {SCA} was inactive in cytosol, and it was activated in vitro by incubation at 30 degrees C. {SCA} was resistant to inhibitors of serine, aspartyl, and metalloproteases, but it was sensitive to N-ethylmaleimide. The enzyme cleaved {SREBP-1} and {SREBP-2} between the Asp and Ser of a conserved sequence {(S/DEPDSP).} The activity was blocked by a tetrapeptide aldehyde, {Ac-Asp-Glu-Ala-Asp-aldehyde} {(Ac-DEAD-CHO).} A purified preparation of {SCA} from hamster liver contained a prominent {20-kDa} polypeptide that could be labeled with {[14C]iodoacetic} acid. Labeling was blocked by {Ac-DEAD-CHO.} Partial amino acid sequence of this polypeptide revealed that it was the hamster equivalent of human {CPP32,} a putative protease whose {cDNA} was recently identified by virtue of sequence homology to {ICE.} {CPP32} and {ICE} have been implicated in apoptosis in animal cells. Whether {SCA/CPP32} participates in vivo in the sterol-regulated activation of {SREBP,} or whether it activates {SREBPs} during apoptosis, remains to be determined.

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  • X. Wang

  • J. T. Pai

  • E. A. Wiedenfeld

  • J. C. Medina

  • C. A. Slaughter

  • J. L. Goldstein

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