G-quadruplexes rescuing protein folding

Abstract

Maintaining the health of the proteome is a critical cellular task. Recently, we found G-quadruplex (G4) nucleic acids are especially potent at preventing protein aggregation in vitro and could at least indirectly improve the protein folding environment of Escherichia coli . However, the roles of G4s in protein folding were not yet explored. Here, through in vitro protein folding experiments, we discover that G4s can accelerate protein folding by rescuing kinetically trapped intermediates to both native and near-native folded states. Time-course folding experiments in E. coli further demonstrate that these G4s primarily improve protein folding quality in E. coli as opposed to preventing protein aggregation. The ability of a short nucleic acid to rescue protein folding opens up the possibility of nucleic acids and ATP-independent chaperones to play considerable roles in dictating the ultimate folding fate of proteins.

An intein‐based biosensor to measure protein stability in vivo

“…Similar to many other fluorescent proteins, TagRFP675 exhibits a slow maturation time (Balleza et al, 2018), resulting in kinetically trapped intermediates and poor folding in E. coli (Son et al, 2023). With the assistance of molecular chaperones, such as GroEL, TagRFP675 can overcome kinetic barriers to fold properly in E. coli (Son et al, 2023). Therefore, we used TagRFP675 to further investigate whether the reporter activity of the intein biosensor is correlated with the folding status of POI in the presence or absence of GroEL (Figure 2).…”

A New Role for RNA G-quadruplexes in Aging and Alzheimer’s Disease

“…As ROS and other protein stresses accumulate with age and according to APOE genotype, rG4s form. Although rG4s can rescue protein folding and could release proteins when unfolded at stress cessation 32 , under persistent stress proteins would not be released from the G-quadruplexes, leading to increased protein oligomerization and aggregation 20 . This function has previously been suggested for RNA in biomolecular condensates more generally 33 .…”

Roles of Nucleic Acids in Protein Folding, Aggregation, and Disease

“…79 Additionally, G-quadruplex-forming sequences assisted in the folding of a protein folding biosensor, TagRFP675 in cells. 80 TagRFP675 does not fold well in E. coli without cooverexpressing a chaperone (such as GroEL or DnaK). Gquadruplex forming sequences had similar protein-folding activity compared to chaperone controls, demonstrating that G-quadruplexes enhanced protein folding in cells.…”

“…79 In vitro assays with chemically denatured TagRFP675 and Gquadruplexes revealed that G-quadruplexes rescued kinetically trapped intermediates to restore TagRFP675 fluorescence. 80 To further explore the mechanism behind the G-quadruplex chaperone activity, two chaperone G-quadruplex sequences with solved NMR structures were systematically mutated. Single-point mutations were able to dramatically affect chaperone activity in either a positive or negative sense.…”

“…Additionally, G-quadruplex-forming sequences assisted in the folding of a protein folding biosensor, TagRFP675 in cells . TagRFP675 does not fold well in E. coli without co-overexpressing a chaperone (such as GroEL or DnaK).…”