title: Distinct properties of wild-type and the amyloidogenic human cystatin C variant of hereditary cerebral hemorrhage with amyloidosis, Icelandic type.
abstract: 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.
authors: Centro Nacional de Biologia Fundamental, Instituto de Salud Carlos III, Madrid, Spain.
authors: M
authors: M
authors: Calero
authors: M
authors: M
authors: Pawlik
authors: C
authors: C
authors: Soto
authors: E M
authors: EM
authors: Castaño
authors: E M
authors: EM
authors: Sigurdsson
authors: A
authors: A
authors: Kumar
authors: G
authors: G
authors: Gallo
authors: B
authors: B
authors: Frangione
authors: E
authors: E
authors: Levy
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D016657:Cerebral Amyloid Angiopathy / Q000235:genetics* / Q000378:metabolism; D002543:Cerebral Hemorrhage / Q000235:genetics* / Q000378:metabolism; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D002645:Chickens; D002942:Circular Dichroism; D003062:Codon / Q000235:genetics; D003198:Computer Simulation; D055316:Cystatin C; D015891:Cystatins / Q000737:chemistry* / Q000235:genetics; D019281:Dimerization; D005973:Glutamine / Q000737:chemistry; D006801:Humans; D007930:Leucine / Q000737:chemistry; D008854:Microscopy, Electron; D008958:Models, Molecular; D020125:Mutation, Missense; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000235:genetics; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D013045:Species Specificity; D017550:Spectroscopy, Fourier Transform Infrared; D014162:Transfection
chemical_list: C525030:CST3 protein, human; D003062:Codon; D055316:Cystatin C; D015891:Cystatins; D009419:Nerve Tissue Proteins; D011993:Recombinant Fusion Proteins; D005973:Glutamine; D007930:Leucine