An Open-faced Chaperonin♦ : Crystal Structures of a Group II Chaperonin Reveal the Open and Closed States Associated with the Protein Folding Cycle
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PROVIDER: S-EPMC2934705 | biostudies-literature | 2010 Sep
REPOSITORIES: biostudies-literature
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Project description:Chaperonins are large protein complexes consisting of two stacked multisubunit rings, which open and close in an ATP-dependent manner to create a protected environment for protein folding. Here, we describe the first crystal structure of a group II chaperonin in an open conformation. We have obtained structures of the archaeal chaperonin from Methanococcus maripaludis in both a peptide acceptor (open) state and a protein folding (closed) state. In contrast with group I chaperonins, in which the equatorial domains share a similar conformation between the open and closed states and the largest motions occurs at the intermediate and apical domains, the three domains of the archaeal chaperonin subunit reorient as a single rigid body. The large rotation observed from the open state to the closed state results in a 65% decrease of the folding chamber volume and creates a highly hydrophilic surface inside the cage. These results suggest a completely distinct closing mechanism in the group II chaperonins as compared with the group I chaperonins.
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Project description:In eukaryotes, CCT is essential for the correct and efficient folding of many cytosolic proteins, most notably actin and tubulin. Structural studies of CCT have been hindered by the failure of standard crystallographic analysis to resolve its eight different subunit types at low resolutions. Here, we exhaustively assess the R value fit of all possible CCT models to available crystallographic data of the closed and open forms with resolutions of 3.8 Å and 5.5 Å, respectively. This unbiased analysis finds the native subunit arrangements with overwhelming significance. The resulting structures provide independent crystallographic proof of the subunit arrangement of CCT and map major asymmetrical features of the particle onto specific subunits. The actin and tubulin substrates both bind around subunit CCT6, which shows other structural anomalies. CCT is thus clearly partitioned, both functionally and evolutionary, into a substrate-binding side that is opposite to the ATP-hydrolyzing side.
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Project description:Mitochondrial Hsp60 (mtHsp60) plays a crucial role in maintaining the proper folding of proteins in the mitochondria. mtHsp60 self-assembles into a ring-shaped heptamer, which can further form a double-ring tetradecamer in the presence of ATP and mtHsp10. However, mtHsp60 tends to dissociate in vitro, unlike its prokaryotic homologue, GroEL. The molecular structure of dissociated mtHsp60 and the mechanism behind its dissociation remain unclear. In this study, we demonstrated that Epinephelus coioides mtHsp60 (EcHsp60) can form a dimeric structure with inactive ATPase activity. The crystal structure of this dimer reveals symmetrical subunit interactions and a rearranged equatorial domain. The α4 helix of each subunit extends and interacts with its adjacent subunit, leading to the disruption of the ATP-binding pocket. Furthermore, an RLK motif in the apical domain contributes to stabilizing the dimeric complex. These structural and biochemical findings provide new insights into the conformational transitions and functional regulation of this ancient chaperonin.
| S-EPMC10053435 | biostudies-literature
Project description:The GroEL-GroES chaperonin system is probably one of the most studied chaperone systems at the level of the molecular mechanism. Since the first reports of a bacterial gene involved in phage morphogenesis in 1972, these proteins have stimulated intensive research for over 40 years. During this time, detailed structural and functional studies have yielded constantly evolving concepts of the chaperonin mechanism of action. Despite of almost three decades of research on this oligomeric protein, certain aspects of its function remain controversial. In this review, we highlight one central aspect of its function, namely, the active intermediates of its reaction cycle, and present how research to this day continues to change our understanding of chaperonin-mediated protein folding.
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Project description:For many enveloped viruses, binding to a receptor(s) on a host cell acts as the first step in a series of events culminating in fusion with the host cell membrane and transfer of genetic material for replication. The envelope glycoprotein (Env) trimer on the surface of HIV is responsible for receptor binding and fusion. Although Env can tolerate a high degree of mutation in five variable regions (V1-V5), and also at N-linked glycosylation sites that contribute roughly half the mass of Env, the functional sites for recognition of receptor CD4 and co-receptor CXCR4/CCR5 are conserved and essential for viral fitness. Soluble SOSIP Env trimers are structural and antigenic mimics of the pre-fusion native, surface-presented Env, and are targets of broadly neutralizing antibodies. Thus, they are attractive immunogens for vaccine development. Here we present high-resolution cryo-electron microscopy structures of subtype B B41 SOSIP Env trimers in complex with CD4 and antibody 17b, or with antibody b12, at resolutions of 3.7 Å and 3.6 Å, respectively. We compare these to cryo-electron microscopy reconstructions of B41 SOSIP Env trimers with no ligand or in complex with either CD4 or the CD4-binding-site antibody PGV04 at 5.6 Å, 5.2 Å and 7.4 Å resolution, respectively. Consequently, we present the most complete description yet, to our knowledge, of the CD4-17b-induced intermediate and provide the molecular basis of the receptor-binding-induced conformational change required for HIV-1 entry into host cells. Both CD4 and b12 induce large, previously uncharacterized conformational rearrangements in the gp41 subunits, and the fusion peptide becomes buried in a newly formed pocket. These structures provide key details on the biological function of the type I viral fusion machine from HIV-1 as well as new templates for inhibitor design.
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Project description:The influenza B M2 (BM2) proton channel is activated by acidic pH to mediate virus uncoating. Unlike influenza A M2 (AM2), which conducts protons with strong inward rectification, BM2 conducts protons both inward and outward. Here we report 1.4- and 1.5-Å solid-state NMR structures of the transmembrane domain of the closed and open BM2 channels in a phospholipid environment. Upon activation, the transmembrane helices increase the tilt angle by 6° and the average pore diameter enlarges by 2.1 Å. BM2 thus undergoes a scissor motion for activation, which differs from the alternating-access motion of AM2. These results indicate that asymmetric proton conduction requires a backbone hinge motion, whereas bidirectional conduction is achieved by a symmetric scissor motion. The proton-selective histidine and gating tryptophan in the open BM2 reorient on the microsecond timescale, similar to AM2, indicating that side chain dynamics are the essential driver of proton shuttling.
| S-EPMC7641042 | biostudies-literature
Project description:Currently, one of the most serious problems in protein-folding simulations for de novo structure prediction is conformational sampling of medium-to-large proteins. In vivo, folding of these proteins is mediated by molecular chaperones. Inspired by the functions of chaperonins, we designed a simple chaperonin-like simulation protocol within the framework of the standard fragment assembly method: in our protocol, the strength of the hydrophobic interaction is periodically modulated to help the protein escape from misfolded structures. We tested this protocol for 38 proteins and found that, using a certain defined criterion of success, our method could successfully predict the native structures of 14 targets, whereas only those of 10 targets were successfully predicted using the standard protocol. In particular, for non-alpha-helical proteins, our method yielded significantly better predictions than the standard approach. This chaperonin-inspired protocol that enhanced de novo structure prediction using folding simulations may, in turn, provide new insights into the working principles underlying the chaperonin system.
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Project description:Escherichia coli glycogen synthase (EcGS, EC 2.4.1.21) is a retaining glycosyltransferase (GT) that transfers glucose from adenosine diphosphate glucose to a glucan chain acceptor with retention of configuration at the anomeric carbon. EcGS belongs to the GT-B structural superfamily. Here we report several EcGS x-ray structures that together shed considerable light on the structure and function of these enzymes. The structure of the wild-type enzyme bound to ADP and glucose revealed a 15.2 degrees overall domain-domain closure and provided for the first time the structure of the catalytically active, closed conformation of a glycogen synthase. The main chain carbonyl group of His-161, Arg-300, and Lys-305 are suggested by the structure to act as critical catalytic residues in the transglycosylation. Glu-377, previously thought to be catalytic is found on the alpha-face of the glucose and plays an electrostatic role in the active site and as a glucose ring locator. This is also consistent with the structure of the EcGS(E377A)-ADP-HEPPSO complex where the glucose moiety is either absent or disordered in the active site.
| S-EPMC2719418 | biostudies-literature
Project description:Thermosomes are group II chaperonins responsible for protein refolding in an ATP-dependent manner. Little is known regarding the conformational changes of thermosomes during their functional cycle due to a lack of high-resolution structure in the open state. Here, we report the first complete crystal structure of thermosome (rATcpn?) in the open state from Acidianus tengchongensis. There is a ?30° rotation of the apical and lid domains compared with the previous closed structure. Besides, the structure reveals a conspicuous hydrophobic patch in the lid domain, and residues locating in this patch are conserved across species. Both the closed and open forms of rATcpn? were also reconstructed by electron microscopy (EM). Structural fitting revealed the detailed conformational change from the open to the closed state. Structural comparison as well as protease K digestion indicated only ATP binding without hydrolysis does not induce chamber closure of thermosome.
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Project description:The GroE chaperonins assist substrate protein (SP) folding by cycling through several conformational states. With each cycle the SP is, in turn, captured, unfolded, briefly encapsulated (t1/2 ∼ 1 s), and released by the chaperonin complex. The protein-folding functional form is the US-football-shaped GroEL:GroES2 complex. We report structures of two such "football" complexes to ∼ 3.7-Å resolution; one is empty whereas the other contains encapsulated SP in both chambers. Although encapsulated SP is not visible on the electron density map, using calibrated FRET and order-of-addition experiments we show that owing to SP-catalyzed ADP/ATP exchange both chambers of the football complex encapsulate SP efficiently only if the binding of SP precedes that of ATP. The two rings of GroEL thus behave as a parallel processing machine, rather than functioning alternately. Compared with the bullet-shaped GroEL:GroES1 complex, the GroEL:GroES2 football complex differs conformationally at the GroEL-GroES interface and also at the interface between the two GroEL rings. We propose that the electrostatic interactions between the ε-NH(3+) of K105 of helix D in one ring with the negatively charged carboxyl oxygen of A109 at the carboxyl end of helix D of the other ring provide the structural basis for negative inter-ring cooperativity.
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