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Transthyretin (TTR) is one of several proteins involved in amyloid disease in humans. Unknown conformational changes of the native state of TTR result in aggregation of TTR molecules into amyloid fibrils, which accumulate in extracellular tissues. This may result in different clinical symptoms, e.g. polyneuropathy or cardiomyopathy, depending on their site of accumulation.

Our long-term goal is to identify structural changes associated with amyloid formation. For this work, structural characterization of TTR from other species than human may provide valuable information.

The three-dimensional X-ray crystallographic structure of TTR from sea bream (Sparus aurata) was determined at 1.75 Å resolution. Human and sea bream TTR were found to be structurally very similar. However, interesting differences were present in the area at and around -strand D, which in fish forms an extended loop region. Interestingly, this area is believed to dissociate from the structure prior to amyloid formation, to allow -strands A and B to participate in polymerization.

During evolution, TTR from different species have developed differences in preference to their natural ligands, the thyroid hormones 3,5,3’-triiodo-L-thyronine (T3) and 3,5,3’,5’-tetraiodo-L-thyronine (T4). While human TTR has higher affinity for T4, the opposite is true in lower vertebrates, e.g. fish and reptiles. We have determined two separate structures of sea bream TTR in complex with T3 and T4, both at 1.9 Å resolution. A significantly wider entrance and narrower thyroid hormone binding channel provide a structural explanation to the differences in thyroid hormone preference between human and piscine TTR.

In a separate work, we identified a novel protein family with structural similarity to TTR, which we named the transthyretin-related protein (TRP) family. To attain information about this protein family, we cloned, expressed, purified and characterized TRP from Escherichia coli (EcTRP). Furthermore, we solved the structure of EcTRP to 1.65 Å resolution. As predicted, EcTRP and human TTR are structurally very similar. Interesting structural differences are found in the area corresponding to the thyroid hormone binding site in TTR, which due to its amino acid conservation within the TRP family we identified as a putative ligand-binding site in TRPs. The function of the TRP is not known, however, recent studies suggest that it might be involved in purine catabolism.

It has been shown that partial acid denaturation of human TTR results in amyloid-fibril formation. Interestingly, we have shown that sea bream TTR also forms amyloid-like fibrils in vitro, even though it shares only 52% sequence identity to human TTR. Corresponding studies on EcTRP did not generate amyloid-like fibrils. EcTRP has 30% sequence identity to human TTR. The fact that two of the proteins form amyloid fibrils and one does not means that they can serve as a model system for the study of amyloid formation. Further studies on these three proteins are currently performed to attain more information about the mechanism of amyloid formation.