This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Transthyretin (TTR) is a plasma carrier protein consisting of 127 amino acid residues. TTR normally exists as a tetramer and binds the hormone thyroxine and the retinol-binding protein-vitamin A complex. Amino acid substitutions in TTR affect the stability of the tetramer and cause the protein to form intermediates that self-associate into amyloid fibrils. Familial transthyretin amyloidosis (ATTR) is associated with the deposition of the TTR variants as amyloid fibrils in tissues and organs. TTR is also associated with senile systemic amyloidosis (SSA), also referred to as senile cardiac amyloidosis (SCA). SSA is a nonhereditary disorder that affects about 25% of individuals over 80 years old. In SSA, the amyloid fibrils are usually composed of wild type TTR and its fragments and are found mainly in the heart. We have found that approximately 85-95% of serum TTR is post-translationally modified by S-sulfonation and S-thiolation (conjugation with cysteine, cysteinyl glycine, and glutathione). Data obtained by ourselves and others indicates that S-Sulfonation of TTR decreases amyloid forming potential and possibly protects against protein aggregation and fibrillogenesis. The extent of thiol conjugation of proteins has been shown to increase with age. The purpose of this study was to evaluate the serum levels of S-sulfonated TTR in SSA patients seen at the BUSM Amyloid Treatment and Research Program, in an attempt to link a biochemical structural feature to disease pathogenesis. We originally hypothesized that increased S-sulfonation of transthyretin (TTR) at Cys10 and interactions with accessory proteins promote fibrillogenesis, and therefore act as effectors of senile systemic amyloidosis (SSA). To test this hypothesis, we analyzed and compared the relative abundance of S-sulfonated TTR isolated from the sera of patients with SSA, primary amyloidosis (AL) with cardiomyopathy, and non-amyloid controls. The results did not indicate that there is a direct link between TTR S-sulfonation and development of SSA. To examine the possibility that the profile of cysteine adducts in serum varies from that in the amyloid deposits, cardiac fibrils were extracted from autopsy tissues obtained on two separate cases of clinically diagnosed SSA. We developed methodology to isolate TTR from these tissues under non-reducing conditions in order to establish the degree of heterogeneity with respect to mass. We observed both truncation, dimerization and Cys-10 modification in the TTR fibrils. Top-down MS analysis enabled assignment of structures to truncated forms of the protein. In 20007, we published these results in Analytical Chemistry. We have carried out stability studies using fluorescently labelled recombinant forms of TTR and its two major metabolites, the S-sulfonated and S-cysteinylated forms and determined by analytical centrifugation that S-sulfonation indeed stabilizes the tetramer, whereas S-cysteinylation destabilizes it. A paper describing the results was recently published in J. Biol. Chem.

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Biotechnology Resource Grants (P41)
Project #
5P41RR010888-13
Application #
7955888
Study Section
Special Emphasis Panel (ZRG1-BCMB-H (40))
Project Start
2009-06-01
Project End
2010-05-31
Budget Start
2009-06-01
Budget End
2010-05-31
Support Year
13
Fiscal Year
2009
Total Cost
$18,853
Indirect Cost
Name
Boston University
Department
Biochemistry
Type
Schools of Medicine
DUNS #
604483045
City
Boston
State
MA
Country
United States
Zip Code
02118
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Srinivasan, Srimathi; Chitalia, Vipul; Meyer, Rosana D et al. (2015) Hypoxia-induced expression of phosducin-like 3 regulates expression of VEGFR-2 and promotes angiogenesis. Angiogenesis 18:449-62
Yu, Xiang; Sargaeva, Nadezda P; Thompson, Christopher J et al. (2015) In-Source Decay Characterization of Isoaspartate and ?-Peptides. Int J Mass Spectrom 390:101-109
Steinhorn, Benjamin S; Loscalzo, Joseph; Michel, Thomas (2015) Nitroglycerin and Nitric Oxide--A Rondo of Themes in Cardiovascular Therapeutics. N Engl J Med 373:277-80

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