Distinct properties of wild-type and the amyloidogenic human cystatin C variant of hereditary cerebral hemorrhage with amyloidosis, Icelandic type.

Variant human cystatin C (L68Q) is an amyloidogenic protein. It deposits in the cerebral vasculature of Icelandic patients with cerebral amyloid angiopathy, leading to stroke. Wild-type and variant cystatin C are cysteine proteinase inhibitors which form concentration dependent inactive dimers; however, variant cystatin C dimerizes at lower concentrations and has an increased susceptibility to a serine protease. We studied the effect of the L68Q amino acid substitution on cystatin C properties, utilizing full length cystatin C purified in mild conditions from media of cells stably transfected with either the wild-type or variant cystatin C genes. The variant cystatin C forms fibrils in vitro detectable by electron microscopy in conditions in which the wild-type protein forms amorphous aggregates. We also show by circular dichroism, steady-state fluorescence and Fourier-transformed infrared spectroscopy that the amino acid substitution modifies cystatin C structure by destabilizing alpha-helical structures and exposing the tryptophan residue to a more polar environment, yielding a more unfolded molecule. These spectral changes demonstrate that variant cystatin C has a three-dimensional structure different from that of the wild-type protein. The structural differences between variant and wild-type cystatin C account for the susceptibility of the variant protein to unfolding, proteolysis and fibrillogenesis.
AuthorsM Calero, M Pawlik, C Soto, E M Castaño, E M Sigurdsson, A Kumar, G Gallo, B Frangione, E Levy
JournalJournal of neurochemistry (J Neurochem) Vol. 77 Issue 2 Pg. 628-37 (Apr 2001) ISSN: 0022-3042 [Print] United States
PMID11299325 (Publication Type: Comparative Study, Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, P.H.S.)
Chemical References
  • CST3 protein, human
  • Codon
  • Cystatin C
  • Cystatins
  • Nerve Tissue Proteins
  • Recombinant Fusion Proteins
  • Glutamine
  • Leucine
  • Amino Acid Substitution
  • Animals
  • Cerebral Amyloid Angiopathy (genetics, metabolism)
  • Cerebral Hemorrhage (genetics, metabolism)
  • Chemistry, Physical
  • Chickens
  • Circular Dichroism
  • Codon (genetics)
  • Computer Simulation
  • Cystatin C
  • Cystatins (chemistry, genetics)
  • Dimerization
  • Glutamine (chemistry)
  • Humans
  • Leucine (chemistry)
  • Microscopy, Electron
  • Models, Molecular
  • Mutation, Missense
  • Nerve Tissue Proteins (chemistry, genetics)
  • Physicochemical Phenomena
  • Protein Conformation
  • Protein Folding
  • Protein Structure, Secondary
  • Recombinant Fusion Proteins (chemistry)
  • Species Specificity
  • Spectroscopy, Fourier Transform Infrared
  • Transfection

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