The kinetics of transthyretin aggregation from acid-unfolded monomers to amyloid fibrils was studied by several biophysical techniques and may be described as a two-step process with transient accumulation of oligomeric species.

Inherited human long-QT2 syndrome (LQTS) results from mutations in the gene encoding the HERG channel. Several LQT2-associated mutations have been mapped to the amino terminal cytoplasmic Per-Arnt-Sim (PAS) domain of the HERG1a channel subunit. Here we have characterized the trafficking properties of some LQT2-associated PAS domain mutants and analyzed rescue of the trafficking mutants by low temperature (27°C) or by the pore blocker drug E4031. We show that the LQT2-associated mutations in the PAS domain of the HERG channel display molecular properties that are distinct from the properties of LQT2-associated mutations in the trans-membrane region. Unlike the latter, many of the tested PAS domain LQT2-associated mutations do not result in trafficking deficiency of the channel. Moreover, the majority of the PAS domain mutations that cause trafficking deficiencies are not rescued by a pore blocking drug. We have also explored the in vitro folding stability properties of isolated mutant PAS domain proteins using a thermal unfolding fluorescence assay and a chemical unfolding assay.

MnTPPS is a metallic water soluble porphyrin with high potential to be used as a contrast agent in photoacoustic tomography. In order to fully understand the interaction between MnTPPS and serum albumin and to investigate the effect of the light induced fast in situ heat deposition by MnTPPS in the protein, we performed several experimental studies using fluorescence and circular dichroism spectroscopies, as well as photoacoustic calorimetry. To identify the possible binding site(s) of the metalloporphyrin in serum albumin and to help interpret the spectroscopic results, a molecular docking exercise was also carried out. The fluorescence data indicate a 1 : 1 stoichiometry for the complex BSA : MnTPPS. The molecular docking results suggest one binding site at the subdomain IB of albumin, where Trp-134 is found, as the main binding site for MnTPPS. The CD data indicate no significant conformational changes of the BSA secondary structure upon MnTPPS binding and even after several minutes of laser excitation of MnTPPS. TR-PAC results show that the in situ heat deposition from MnTPPS does not cause any significant transient conformational change to the BSA structure. In conclusion, this work demonstrates that MnTPPS, in addition to the necessary physical and chemical properties to be used as a contrast agent in photoacoustic tomography, can be effectively carried by albumin and that in situ heat release following light absorption does not cause any significant damage to the protein structure.