Characterization of Intrinsically Disordered Prostate Associated Gene (PAGE5) at Single Residue Resolution by NMR Spectroscopy

Abstract

BackgroundThe Cancer-Testis antigens (CTA) are proteins expressed in human germ line and certain cancer cells. CTAs form a large gene family, representing 10% of X-chromosomal genes. They have high potential for cancer-specific immunotherapy. However, their biological functions are currently unknown. Prostate associated genes (PAGE) are characterized as CTAs. PAGE5 is one of six proteins belonging to this protein family, also called CT16.Methodology/Principal findingsIn this study we show, using bioinformatics, chromatographic and solution state NMR spectroscopic methods, that PAGE5 is an intrinsically disordered protein (IDP).Conclusion/SignificanceThe study stands out as the first time structural characterization of the PAGE family protein and introduces how solution state NMR spectroscopy can be effectively utilized for identification of molecular recognition regions (MoRF) in IDPs, known often as transiently populated secondary structures.

Phosphorylation-induced Conformational Ensemble Switching in an Intrinsically Disordered Cancer/Testis Antigen

“…Comparison with Other PAGE Family Members-The only other cancer/testis antigen studied in detail using biophysical methods is PAGE5 (29). However, the role of posttranslational modifications in PAGE5 remains unknown.…”

“…Transient ␤-extended and helical structures are represented by arrows and cylinders, respectively. PAGE5 preferences from an earlier study (29) are shown in white at the bottom of the alignment.…”

“…To our knowledge, this is the first detailed study on the effects of phosphorylation in an intrinsically disordered CT-X antigen family member that plays an important role in the diseased prostate. Only one other completely disordered CT-X antigen has been investigated from a biophysical perspective to date, a splice variant of PAGE5 that lacks part of the N terminus (29). However, the biological function(s) of PAGE5 in the prostate and the effect of posttranslational modifications on its structure and function remain unknown.…”

Human Adenosine A2A Receptor Binds Calmodulin with High Affinity in a Calcium-Dependent Manner

“…3 A). The spectrum shows poorly dispersed crosspeaks, with the proton chemical shifts ranging from 7.5 to 8.5 1 H ppm, indicating that A2A-ct undergoes fast conformational dynamics in solution, which is typical for highly disordered proteins (69). The conformational heterogeneity of intrinsically disordered proteins calls for special means for the NMR signal assignment.…”

“…NMR spectroscopy can be employed to study structural features of disordered proteins at single-residue resolution (69). To confirm the disordered nature of A2A-ctL in solution, we measured a two-dimensional 1 H, 15 N-HSQC spectrum of A2A-ctL at 800 MHz 1 H frequency (Fig.…”