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Physico-chemical properties of the N-terminally truncated L68Q cystatin C found in amyloid deposits of brain haemorrhage patients.

Gerhartz, Bernd and Abrahamson, Magnus LU (2002) In Biological Chemistry 383(2). p.301-305
Abstract
Cystatin C, a major extracellular cysteine proteinase inhibitor, is deposited as amyloid in brain haemorrhage patients with hereditary cystatin C amyloid angiopathy (HCCAA). A disease-causing mutation on the genetic level results in the substitution Leu68-->Gln (L68Q) in cystatin C, which causes protein instability. Besides carrying the L68Q substitution, cystatin C in amyloid deposits isolated from patients is N-terminally truncated by 10 amino acids. To elucidate the role of the N-terminal truncation for protein stability and aggregation properties, (delta1-10,L68Q)-cystatin C was produced in an Escherichia coli expression system and characterised. Unlike wild-type cystatin C, this variant rapidly dimerised under physiological... (More)
Cystatin C, a major extracellular cysteine proteinase inhibitor, is deposited as amyloid in brain haemorrhage patients with hereditary cystatin C amyloid angiopathy (HCCAA). A disease-causing mutation on the genetic level results in the substitution Leu68-->Gln (L68Q) in cystatin C, which causes protein instability. Besides carrying the L68Q substitution, cystatin C in amyloid deposits isolated from patients is N-terminally truncated by 10 amino acids. To elucidate the role of the N-terminal truncation for protein stability and aggregation properties, (delta1-10,L68Q)-cystatin C was produced in an Escherichia coli expression system and characterised. Unlike wild-type cystatin C, this variant rapidly dimerised under physiological conditions. Two unfolding intermediates of (delta1-10,L68Q)-cystatin C were identified, under the same pH and ionic strength conditions as required to form intermediates of full-length L68Q cystatin C. No evidence was found that the N-terminal truncation per se alters protein stability and leads to higher forms of aggregation. Monomeric as well as dimeric L68Q cystatin C incubated with neutrophil elastase was truncated as in HCCAA patients' amyloid. A protein variant with a thrombin cleavage site placed in front of residue Gly11 in L68Q cystatin C was constructed and used to confirm that the N-terminal segment is similarly accessible to proteinases in the monomeric and dimeric states of L68Q cystatin C. Thus, the N-terminal segment of L68Q cystatin C is exposed to proteolytic attack and does not seem to be involved in intramolecular contacts leading to dimerisation or higher-order aggregation. We conclude that the N-terminal truncation likely is an event secondary to amyloid formation, and of no relevance for the development of HCCAA. (Less)
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author
and
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Biological Chemistry
volume
383
issue
2
pages
301 - 305
publisher
De Gruyter
external identifiers
  • pmid:11934268
  • wos:000174545300008
  • scopus:0036118589
ISSN
1437-4315
language
English
LU publication?
yes
id
c5ca64c6-e232-42ef-ade8-a7ee67864876 (old id 107405)
alternative location
http://www.degruyter.de/journals/bc/272_3065_ENU_h.htm
date added to LUP
2016-04-01 11:51:01
date last changed
2022-01-26 19:09:07
@article{c5ca64c6-e232-42ef-ade8-a7ee67864876,
  abstract     = {{Cystatin C, a major extracellular cysteine proteinase inhibitor, is deposited as amyloid in brain haemorrhage patients with hereditary cystatin C amyloid angiopathy (HCCAA). A disease-causing mutation on the genetic level results in the substitution Leu68-->Gln (L68Q) in cystatin C, which causes protein instability. Besides carrying the L68Q substitution, cystatin C in amyloid deposits isolated from patients is N-terminally truncated by 10 amino acids. To elucidate the role of the N-terminal truncation for protein stability and aggregation properties, (delta1-10,L68Q)-cystatin C was produced in an Escherichia coli expression system and characterised. Unlike wild-type cystatin C, this variant rapidly dimerised under physiological conditions. Two unfolding intermediates of (delta1-10,L68Q)-cystatin C were identified, under the same pH and ionic strength conditions as required to form intermediates of full-length L68Q cystatin C. No evidence was found that the N-terminal truncation per se alters protein stability and leads to higher forms of aggregation. Monomeric as well as dimeric L68Q cystatin C incubated with neutrophil elastase was truncated as in HCCAA patients' amyloid. A protein variant with a thrombin cleavage site placed in front of residue Gly11 in L68Q cystatin C was constructed and used to confirm that the N-terminal segment is similarly accessible to proteinases in the monomeric and dimeric states of L68Q cystatin C. Thus, the N-terminal segment of L68Q cystatin C is exposed to proteolytic attack and does not seem to be involved in intramolecular contacts leading to dimerisation or higher-order aggregation. We conclude that the N-terminal truncation likely is an event secondary to amyloid formation, and of no relevance for the development of HCCAA.}},
  author       = {{Gerhartz, Bernd and Abrahamson, Magnus}},
  issn         = {{1437-4315}},
  language     = {{eng}},
  number       = {{2}},
  pages        = {{301--305}},
  publisher    = {{De Gruyter}},
  series       = {{Biological Chemistry}},
  title        = {{Physico-chemical properties of the N-terminally truncated L68Q cystatin C found in amyloid deposits of brain haemorrhage patients.}},
  url          = {{http://www.degruyter.de/journals/bc/272_3065_ENU_h.htm}},
  volume       = {{383}},
  year         = {{2002}},
}