Title
Interplay between Structure and Charge as a Key to Allosteric Modulation of Human 20S Proteasome by the Basic Fragment of HIV-1 Tat Protein
Author
Przemysław Karpowicz, Paweł A. Osmulski, Julia Witkowska, Emilia Sikorska, Małgorzata Giżyńska, Agnieszka Belczyk-Ciesielska, Maria E. Gaczynska, Elżbieta Jankowska
Year
2015
Journal
PLoS ONE
Abstract
The proteasome is a giant protease responsible for degradation of the majority of cytosolic proteins. Competitive inhibitors of the proteasome are used against aggressive blood cancers. However, broadening the use of proteasome-targeting drugs requires new mechanistic approaches to the enzyme’s inhibition. In our previous studies we described Tat1 peptide, an allosteric inhibitor of the proteasome derived from a fragment of the basic domain of HIV-Tat1 protein. Here, we attempted to dissect the structural determinants of the proteasome inhibition by Tat1. Single- and multiple- alanine walking scans were performed. Tat1 analogs with stabilized beta-turn conformation at positions 4–5 and 8–9, pointed out by the molecular dynamics modeling and the alanine scan, were synthesized. Structure of Tat1 analogs were analyzed by circular dichroism, Fourier transform infrared and nuclear magnetic resonance spectroscopy studies, supplemented by molecular dynamics simulations. Biological activity tests and structural studies revealed that high flexibility and exposed positive charge are hallmarks of Tat1 peptide. Interestingly, stabilization of a beta-turn at the 8–9 position was necessary to significantly improve the inhibitory potency.
Instrument
J-815
Keywords
Circular dichroism, Secondary structure, Biochemistry