Identification and characterization of a human transthyretin variant

An apparent Mr variant of plasma transthyretin (TTR), previously detected using 2-D PAGE, is the first reported occurrence of this type of human TTR variant. We characterized the variant TTR to determine the nature of this difference. Comparative tryptic peptide maps of variant and normal TTR and sequencing of peptides which differed indicated the variant contained a single amino acid substitution of valine for tyrosine at position 116. Because such a change requires two nucleotide substitutions, we postulate the variant arose through mutation in codon 116 of a heretofore unrecognized polymorphic or rare variant allele of TTR.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/26538/1/0000077.pd

Variation in amount of wild-type transthyretin in different fibril and tissue types in ATTR amyloidosis

Familial transthyretin (TTR) amyloidosis is caused by a mutation in the TTR gene, although wild-type (wt) TTR is also incorporated into the amyloid fibrils. Liver transplantation (LT) is the prevailing treatment of the disease and is performed in order to eliminate the mutant TTR from plasma. The outcome of the procedure is varied; especially problematic is a progressive cardiomyopathy seen in some patients, presumably caused by continued incorporation of wtTTR. What determines the discrepancy in outcome is not clear. We have previously shown that two structurally distinct amyloid fibrils (with or without fragmented ATTR) are found among ATTRV30M patients. In this study, we investigated the proportion of wtATTR in cardiac and adipose amyloid from patients having either fibril type. It was found that cardiac amyloid more easily incorporates wtTTR than adipose amyloid, offering a potential explanation for the vulnerability of cardiac tissue for continued amyloidosis after LT. In cardiac tissue, fibrils with fragmented ATTR contained a higher wt proportion than fibrils without, suggesting that continued incorporation of wtTTR after LT, perhaps, can take place more easily in these patients. In adipose tissue, a rapid increase in wt proportion after LT indicates that a rather fast turnover of the deposits must occur. A difference in wt proportion between the fibril types was seen post-LT but not pre-LT, possibly caused by differences in turnover rate. Conclusively, this study further establishes the basic dissimilarities between the two fibril types and demonstrates that their role in LT outcome needs to be further investigated

Comparative calorimetric study of non-amyloidogenic and amyloidogenic variants of the homotetrameric protein transthyretin

Familial amyloidotic polyneuropathy (FAP) is an autosomal dominant hereditary type of amyloidosis involving amino acid substitutions in transthyretin (TTR). V30M-TTR is the most frequent variant, and L55P-TTR is the variant associated with the most aggressive form of FAP. The thermal stability of the wild-type, V30M-TTR, L55P-TTR and a non-amyloidogenic variant, T119M-TTR, was studied by high-sensitivity differential scanning calorimetry (DSC). The thermal unfolding of TTR is a spontaneous reversible process involving a highly co-operative transition between folded tetramers and unfolded monomers. All variants of transthyretin are very stable to the thermal unfolding that occurs at very high temperatures, most probably because of their oligomeric structure. The data presented in this work indicated that for the homotetrameric form of the wild-type TTR and its variants, the order of stability is as follows: wild-type TTRT119M-TTR>L55P-TTR>V30M-TTR, which does not correlate with their known amyloidogenic potential.http://www.sciencedirect.com/science/article/B6TFB-41NK91T-5/1/472cf0fa1a8c059202a368bff378672

Structural Confirmation of a Rare Isocyanoterpene: Synthesis of Isoneoamphilectane and its C-7 and C-8 Epimers

This dissertation describes our efforts towards a total synthesis of the rare isocyanoterpene, isoneoamphilectane. Chapter 1 includes an introduction to the amphilectane family of natural products along with a discussion on characterization, biosynthesis, biological relevance, and synthetic challenges. Chapter 2 describes our initial synthetic efforts aimed at applying an intramolecular trimethylenemethane [3+2] cycloaddition to construct the isoneoamphilectane core. Chapter 3 details a different strategy wherein we pursued a Michael addition/enolate alkylation approach to the tricyclic core of isoneoamphilectane. Chapter 4 discusses our hypothesis for a potential structural revision, which guided our synthetic design and motivated the synthesis of two diastereomers of the reported natural product: 7,8-di-epi-isoneoamphilectane and 8-epi-isoneoamphilectane. With greater insight into the natural product’s stereochemical assignment, we embarked on a synthesis of isoneoamphilectane hinging on a challenging cis-to-trans decalone epimerization. In Chapter 5, we describe our investigations into contrathermodynamic epimerization strategies with the discovery of a cyclic sulfite-based pinacol-like rearrangement to install the strained ring system. Additional key steps in our synthesis include an acyclic Mukaiyama–Michael addition, an intramolecular alkoxide-directed elimination, and an HAT-mediated alkene hydroazidation. In conclusion, we report the first total synthesis of isoneoamphilectane in 23 steps from known materials