Protein folding and aggregation in bacteria

Abstract

Proteins might experience many conformational changes and interactions during their lifetimes, from their synthesis at ribosomes to their controlled degradation. Because, in most cases, only folded proteins are functional, protein folding in bacteria is tightly controlled genetically, transcriptionally, and at the protein sequence level. In addition, important cellular machinery assists the folding of polypeptides to avoid misfolding and ensure the attainment of functional structures. When these redundant protective strategies are overcome, misfolded polypeptides are recruited into insoluble inclusion bodies. The protein embedded in these intracellular deposits might display different conformations including functional and beta-sheet-rich structures. The latter assemblies are similar to the amyloid fibrils characteristic of several human neurodegenerative diseases. Interestingly, bacteria exploit the same structural principles for functional properties such as adhesion or cytotoxicity. Overall, this review illustrates how prokaryotic organisms might provide the bedrock on which to understand the complexity of protein folding and aggregation in the cell.

Fig. 1

Vectorial-assisted folding of a newly synthesized polypeptide involving the three major chaperone systems in the bacterial cytosol. ELS complex reproduced with permission from Ref. [89]

Fig. 2

Structure and function of the GroEL–GroES (ELS) complex. a Top view of the complex looking down from the GroES-binding (cis) side. b Side view, trans GroEL ring is shown in red, cis GroEL ring in green and GroES in gold. c Each subunit in each GroEL ring is composed of three domains: the apical, equatorial, and intermediate domains. Misfolded polypeptides bind to the apical domain (in red), the equatorial domain (in blue) binds and hydrolyzes ATP and the intermediate domain (in green) acts as connector. GroES binding to GroEL-ATP causes the apical domain to move upward and rotate 120°. This large conformational change increases the size of central cavity where the polypeptide resides. Accordingly, the cis and trans rings display different volumes [81]. d cis mechanism of ELS complex. (I) The GroEL apical domains in the open the ring capture misfolded polypeptides. (II) Formation the GroEL-ATP-GroES complex promotes conformational changes in the apical domain resulting in substrate release inside the cis cavity. (III) Folding continues until ATP hydrolysis results in debilitation of the GroEL–GroES interaction. (IV) ATP binding to the trans-ring favors the release of substrate from the cis-ring. Consecutive cycles can be necessary until a correct protein structure is attained. (V) Binding of GroES to GroEL allows the rings to alternate between binding-active and folding-active states. e trans mechanism of ELS. (I) The apical domains in the open ring of GroEL–GroES-ADP complex can capture misfolded polypeptides. (II) Binding occurs through hydrophobic interactions. (III) Binding of ATP to the cis-equatorial domain of GroEL promotes the release of GroES and ADP from the trans ring. (IV) ATP and GroES rebind to the trans GroEL ring, promoting a conformational change on the cis GroEL ring and liberating the polypeptide into the solution. The committed form of the polypeptide is not perfectly folded but will become fully active as it reaches the solution. Protein conformers unable to attain the native state can rebind to GroEL and go through subsequent cycles. a–c adapted with permission from Ref. [89]; d adapted with permission from Ref. [81]; and e adapted with permission from Ref. [97]

Fig. 3

Localization of IBs to the cellular poles. a Schematic diagram illustrating IB segregation within a bacterial population. Independently of the initial position in the mother cell, at polar, mid-cell, or quarter-cell positions, after a reduced number of cellular divisions, IBs would become invariably located and consistently inherited by the old-pole cell (in red) whereas new poles (in blue) and daughter cells become free of aggregates. b, c Migration of protein aggregates to the bacterial poles. b Images of a time course experiment indicating the location of a recombinant aggregation-prone protein fused to GFP after induction of its expression. c Next to each frame is shown the frequency of GFP fluorescence at each particular point over time according to the scale on the right, where brighter colors indicate higher GFP permanence. a Adapted with permission from Ref. [25]; b, c adapted with permission from Ref. [26]

Fig. 4

Remodeling of bacterial protein aggregates. a Structural model of ClpB hexamer with bound ATP. An individual subunit is shown in blue and ATP in red. b Disaggregation of bacterial aggregates by ClpB in combination with the DnaK system. The DnaK chaperone system interacts with ClpB, acting before or together with ClpB. DnaK is likely involved in the presentation of substrates to ClpB (i) and/or in the hydrolysis of ATP. ClpB acts as the main disaggregase. It extracts proteins from the aggregate by translocating them through their central channel (ii) and releasing them in an unfolded form into the solution (iii) where they can refold spontaneously or with assistance of other chaperones including the DnaK system (iv). a Reproduced from [125] and b adapted with permission from [125]

Fig. 5

Conformations adopted by a recombinant protein inside bacteria. After protein synthesis, the polypeptide can fold, either spontaneously or with the assistance of chaperones, or remain totally or partially unfolded. These initially soluble structures can aggregate, preferably through selective interactions, to form a supramolecular IB in which multiple conformations can coexist. The protein quality machinery acts on both the soluble and insoluble protein conformers, promoting a kinetic equilibrium between both cellular fractions. These actions result in functional and inactive protein structures coexisting in both the soluble and insoluble compartments. Accordingly, solubility alone does not present an accurate measure of protein quality during recombinant expression and IBs cannot be excluded as a source of functional protein

Fig. 6

Formation of functional amyloid-like structures allows streptomycetes to invade the air. The water surface tension at the substrate-air interface is reduced by proteins that self-assemble into an amphipathic rigid membrane. This membrane is composed of amyloid-like fibrils formed by the supramolecular organization of chaplins. Aerial hyphae continue to secrete these proteins that assemble at the hyphal surface, making them hydrophobic. Figure adapted with permission from Ref. [178]