Spectroscopic studies reveal conformational flexibility of intrinsically unstructured protein HYPK

The chaperone-like huntingtin-interacting protein, HYPK, has unusual biophysical behavior like an intrinsically unstructured protein (IUP). The protein exists as a (pre-) molten globule with ~37% residual structure and shows com- paction in presence of Ca++. HYPK contains no intrinsic fluorophore other than a single tyrosine and displays an anomalous fluorescence peak at around 340 nm. The anomalous peak is re- duced to 303 nm by the addition of guanidine hydrochloride and at low pH, concomitant with the emission spectrum of L-tyrosine. At high pH the peak is shifted to ~350 nm with a reduction in intensity. In presence of sodium perchlorate there is no shift in HYPK fluorescence emission peak from ~340 nm suggesting localization of the lone tyrosine residue in helical regions. In CD experiments, however, a shift in local secondary structure is noticed upon perchlorate treatment. Acrylamide quenching experiments at different Ca++ concentrations demonstrate that Ca++ does not alter the accessibility of the tyro- sine to acrylamide. In the absence of any tryp- tophan contamination, these observations vali- date that, in vitro, HYPK possesses a loosely associated (pre-) molten globule like conforma- tion with the lone tyrosine being situated within an α-helix.

Novel Adaptors of Amyloid Precursor Protein Intracellular Domain and Their Functional Implications

Amyloid precursor protein intracellular domain （AICD） is one of the potential candidates in deciphering the complexity of Alzheimer＇s disease. It plays important roles in determining cell fate and neurodegeneration through its interactions with several adaptors. The pres- ence or absence of phosphorylation at specific sites determines the choice of partners. In this study, we identified 20 novel AICD- interacting proteins by in vitro pull down experiments followed by 2D gel electrophoresis and MALDI-MS analysis. The identified proteins can be grouped into different functional classes including molecular chaperones, structural proteins, signaling and transport molecules, adaptors, motor proteins and apoptosis determinants. Interactions of nine proteins were further validated either by colocal- ization using confocal imaging or by co-immunoprecipitation followed by immunoblotting. The cellular functions of most of the proteins can be correlated with AD. Hence, illustration of their interactions with AICD may shed some light on the disease pathophysiology.