Spectroscopic studies reveal conformational flexibility of intrinsically unstructured protein HYPK

Swasti Raychaudhuri,Shreoshi Palchoudhuri,Kamalika Roy Choudhury,Shradha Chopra,Debashis Mukhopadhyay,Nitai P Bhattacharyya

doi:10.4236/jbpc.2011.24051

Swasti Raychaudhuri, Shreoshi Palchoudhuri + Show 4 more

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https://doi.org/10.4236/jbpc.2011.24051

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Abstract

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.

Highlights

Huntingtin Yeast-two hybrid Protein K (HYPK) is a#These authors contributed to this work.huntingtin-interacting protein that has been shown to reduce aggregation and apoptosis in a Huntington’s disease (HD) model [1]
HYPK has a wide fluorescence emission peak encompassing 330 - 350 nm compared to L-tyrosine which has a distinct peak at 303 nm (Figure 1(a))
There was no marked change in absorption and fluorescence spectra of HYPK with these treatments suggesting the absence of contamination

Summary

Introduction

Huntingtin Yeast-two hybrid Protein K (HYPK) is a#These authors contributed to this work.huntingtin-interacting protein that has been shown to reduce aggregation and apoptosis in a Huntington’s disease (HD) model [1]. Our previous study showed that HYPK is an intrinsically unstructured protein (IUP) with a pre-molten globule like conformation [4]. It is hypothesized that IUPs interact with multiple partners by attaining different conformations in variable biological contexts [5,6]. Disorder-to-order transition is a characteristic of intrinsic disordered protein upon binding with different partners. It has been reported recently that the anchor residues in the disordered proteins are an important factor in controlling the specific interaction between IDPs and their targets and play vital role in stabilizing the transient binding complexes [8]. IUPs seem to play important roles in evolution [9] and the binding promiscuity of intrinsically disordered protein plays major roles in the diverse functional properties of unstructured protein

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