Protein folding while chaperone bound is dependent on weak interactions

Abstract

It is generally assumed that protein clients fold following their release from chaperones instead of folding while remaining chaperone-bound, in part because binding is assumed to constrain the mobility of bound clients. Previously, we made the surprising observation that the ATP-independent chaperone Spy allows its client protein Im7 to fold into the native state while continuously bound to the chaperone. Spy apparently permits sufficient client mobility to allow folding to occur while chaperone bound. Here, we show that strengthening the interaction between Spy and a recently discovered client SH3 strongly inhibits the ability of the client to fold while chaperone bound. The more tightly Spy binds to its client, the more it slows the folding rate of the bound client. Efficient chaperone-mediated folding while bound appears to represent an evolutionary balance between interactions of sufficient strength to mediate folding and interactions that are too tight, which tend to inhibit folding.

Mechanism of the small ATP-independent chaperone Spy is substrate specific

“…In this mechanism, substrate release is not a prerequisite for substrate folding. We have found that the folding-while-bound mechanism is dependent on relatively weak chaperone–substrate interactions 9 . Variants of Spy that bind SH3 with stronger affinity than does wild-type (WT) Spy significantly slow the folding of bound substrate 9 .…”

“…We have found that the folding-while-bound mechanism is dependent on relatively weak chaperone–substrate interactions 9 . Variants of Spy that bind SH3 with stronger affinity than does wild-type (WT) Spy significantly slow the folding of bound substrate 9 . The substrates of Spy tested so far have been small, topologically simple, all β-sheet or all α-helical model proteins.…”

“…Substrates bound to ATP-independent chaperones such as trigger factor, SecB, and the sHsps can adopt a wide range of conformations ranging from near-native to unfolded states 5 – 7 . We have shown that the ATP-independent chaperone Spy not only binds protein folding intermediates of substrate proteins such as Im7 and SH3 but also allows for the folding of substrate proteins while they remain chaperone bound 8 , 9 . Spy loosely binds to different folding states of these model substrate proteins.…”

Características comunes de las chaperonas pequeñas y diméricas

“…En la región cóncava es donde interaccionan las proteínas sustrato. Esta región tiene cuatro zonas hidrofóbicas rodeadas de aminoácidos hidrofílicos con carga positiva, pertenecientes a la región N-terminal flexible y al asa que conecta H1 con H2 (Koldewey et al, 2017;Kwon et al, 2010;Wu, Stull, Lee & Bardwell, 2019)…”

“…La proteína Spy ha sido muy estudiada y sirve como modelo de la interacción chaperona/proteína sustrato. Esto se debe al tamaño pequeño de Spy y a que se han identificado algunos de sus sustratos específicos, como la proteína Im7 (Wu et al, 2019). El mecanismo por el que Spy favorece el plegamiento de otras proteínas se debe a que la proteína sustrato se une a la chaperona a partir de interacciones electrostáticas transitorias de largo alcance.…”