Cryoprotective activities of FK20, a human genome-derived intrinsically disordered peptide against cryosensitive enzymes without a stereospecific molecular interaction

Abstract

Background Intrinsically disordered proteins (IDPs) have been shown to exhibit cryoprotective activity toward other cellular enzymes without any obvious conserved sequence motifs. This study investigated relationships between the physical properties of several human genome-derived IDPs and their cryoprotective activities. Methods Cryoprotective activity of three human-genome derived IDPs and their truncated peptides toward lactate dehydrogenase (LDH) and glutathione S-transferase (GST) was examined. After the shortest cryoprotective peptide was defined (named FK20), cryoprotective activity of all-D-enantiomeric isoform of FK20 (FK20-D) as well as a racemic mixture of FK20 and FK20-D was examined. In order to examine the lack of increase of thermal stability of the target enzyme, the CD spectra of GST and LDH in the presence of a racemic mixture of FK20 and FK20-D at varying temperatures were measured and used to estimate Tm. Results Cryoprotective activity of IDPs longer than 20 amino acids was nearly independent of the amino acid length. The shortest IDP-derived 20 amino acid length peptide with sufficient cryoprotective activity was developed from a series of TNFRSF11B fragments (named FK20). FK20, FK20-D, and an equimolar mixture of FK20 and FK20-D also showed similar cryoprotective activity toward LDH and GST. Tm of GST in the presence and absence of an equimolar mixture of FK20 and FK20-D are similar, suggesting that IDPs’ cryoprotection mechanism seems partly from a molecular shielding effect rather than a direct interaction with the target enzymes.

Keywords

• Intrinsically disordered proteins
• Cryoprotection
• Thermostability
• all-D-enantiomeric peptide
• Glutathione-S-transferase
• Molecular shield