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1.
Signal recognition particle (SRP), together with its receptor (SR), mediates the targeting of ribosome-nascent chain complexes to the endoplasmic reticulum. Using protein cross-linking, we detected distinct modes in the binding of SRP to the ribosome. During signal peptide recognition, SRP54 is positioned at the exit site close to ribosomal proteins L23a and L35. When SRP54 contacts SR, SRP54 is rearranged such that it is no longer close to L23a. This repositioning may allow the translocon to dock with the ribosome, leading to insertion of the signal peptide into the translocation channel. 相似文献
2.
Ataide SF Schmitz N Shen K Ke A Shan SO Doudna JA Ban N 《Science (New York, N.Y.)》2011,331(6019):881-886
Cotranslational targeting of membrane and secretory proteins is mediated by the universally conserved signal recognition particle (SRP). Together with its receptor (SR), SRP mediates the guanine triphosphate (GTP)-dependent delivery of translating ribosomes bearing signal sequences to translocons on the target membrane. Here, we present the crystal structure of the SRP:SR complex at 3.9 angstrom resolution and biochemical data revealing that the activated SRP:SR guanine triphosphatase (GTPase) complex binds the distal end of the SRP hairpin RNA where GTP hydrolysis is stimulated. Combined with previous findings, these results suggest that the SRP:SR GTPase complex initially assembles at the tetraloop end of the SRP RNA and then relocalizes to the opposite end of the RNA. This rearrangement provides a mechanism for coupling GTP hydrolysis to the handover of cargo to the translocon. 相似文献
3.
Two structurally homologous guanosine triphosphatase (GTPase) domains interact directly during signal recognition particle (SRP)-mediated cotranslational targeting of proteins to the membrane. The 2.05 angstrom structure of a complex of the NG GTPase domains of Ffh and FtsY reveals a remarkably symmetric heterodimer sequestering a composite active site that contains two bound nucleotides. The structure explains the coordinate activation of the two GTPases. Conformational changes coupled to formation of their extensive interface may function allosterically to signal formation of the targeting complex to the signal-sequence binding site and the translocon. We propose that the complex represents a molecular "latch" and that its disengagement is regulated by completion of assembly of the GTPase active site. 相似文献
4.
Peluso P Herschlag D Nock S Freymann DM Johnson AE Walter P 《Science (New York, N.Y.)》2000,288(5471):1640-1643
The mechanism by which a signal recognition particle (SRP) and its receptor mediate protein targeting to the endoplasmic reticulum or to the bacterial plasma membrane is evolutionarily conserved. In Escherichia coli, this reaction is mediated by the Ffh/4.5S RNA ribonucleoprotein complex (Ffh/4.5S RNP; the SRP) and the FtsY protein (the SRP receptor). We have quantified the effects of 4.5S RNA on Ffh-FtsY complex formation by monitoring changes in tryptophan fluorescence. Surprisingly, 4.5S RNA facilitates both assembly and disassembly of the Ffh-FtsY complex to a similar extent. These results provide an example of an RNA molecule facilitating protein-protein interactions in a catalytic fashion. 相似文献
5.
Requirement of GTP hydrolysis for dissociation of the signal recognition particle from its receptor. 总被引：23，自引：0，他引：23
The signal recognition particle (SRP) directs signal sequence specific targeting of ribosomes to the rough endoplasmic reticulum. Displacement of the SRP from the signal sequence of a nascent polypeptide is a guanosine triphosphate (GTP)-dependent reaction mediated by the membrane-bound SRP receptor. A nonhydrolyzable GTP analog can replace GTP in the signal sequence displacement reaction, but the SRP then fails to dissociate from the membrane. Complexes of the SRP with its receptor containing the nonhydrolyzable analog are incompetent for subsequent rounds of protein translocation. Thus, vectorial targeting of ribosomes to the endoplasmic reticulum is controlled by a GTP hydrolysis cycle that regulates the affinity between the SRP, signal sequences, and the SRP receptor. 相似文献
6.
The signal recognition particle (SRP) is a universally conserved ribonucleoprotein complex that mediates the cotranslational targeting of secretory and membrane proteins to cellular membranes. A crucial early step in SRP assembly in archaea and eukarya is the binding of protein SRP19 to specific sites on SRP RNA. Here we report the 1.8 angstrom resolution crystal structure of human SRP19 in complex with its primary binding site on helix 6 of SRP RNA, which consists of a stem-loop structure closed by an unusual GGAG tetraloop. Protein-RNA interactions are mediated by the specific recognition of a widened major groove and the tetraloop without any direct protein-base contacts and include a complex network of highly ordered water molecules. A model of the assembly of the SRP core comprising SRP19, SRP54, and SRP RNA based on crystallographic and biochemical data is proposed. 相似文献
7.
利用同源建模得到杀虫晶体蛋白Cry1Ac的三维结构,之后模拟其与配体N-已酰半乳糖胺的对接,预测关键的氨基酸残基:N482、Q506、S501、L505和V483。Cry1Ac的虚拟突变体与N-已酰半乳糖胺之间的分子对接分析结果表明,R466、R468、R470和R472可能对维持Cry1Ac的DomainⅢ稳定构象起着重要作用。N482和Q506两个残基均含有酰氨基,在对接中易于形成多个氢键,这对稳定对接具有重要作用。研究结果可提供一些有用的信息,用于指导杀虫晶体蛋白的理性设计。 相似文献
8.
Eubacteria inactivate their ribosomes as 100S dimers or 70S monomers upon entry into stationary phase. In Escherichia coli, 100S dimer formation is mediated by ribosome modulation factor (RMF) and hibernation promoting factor (HPF), or alternatively, the YfiA protein inactivates ribosomes as 70S monomers. Here, we present high-resolution crystal structures of the Thermus thermophilus 70S ribosome in complex with each of these stationary-phase factors. The binding site of RMF overlaps with that of the messenger RNA (mRNA) Shine-Dalgarno sequence, which prevents the interaction between the mRNA and the 16S ribosomal RNA. The nearly identical binding sites of HPF and YfiA overlap with those of the mRNA, transfer RNA, and initiation factors, which prevents translation initiation. The binding of RMF and HPF, but not YfiA, to the ribosome induces a conformational change of the 30S head domain that promotes 100S dimer formation. 相似文献
9.
Stengel KF Holdermann I Cain P Robinson C Wild K Sinning I 《Science (New York, N.Y.)》2008,321(5886):253-256
Secretory and membrane proteins carry amino-terminal signal sequences that, in cotranslational targeting, are recognized by the signal recognition particle protein SRP54 without sequence specificity. The most abundant membrane proteins on Earth are the light-harvesting chlorophyll a/b binding proteins (LHCPs). They are synthesized in the cytoplasm, imported into the chloroplast, and posttranslationally targeted to the thylakoid membrane by cpSRP, a heterodimer formed by cpSRP54 and cpSRP43. We present the 1.5 angstrom crystal structure of cpSRP43 characterized by a unique arrangement of chromodomains and ankyrin repeats. The overall shape and charge distribution of cpSRP43 resembles the SRP RNA, which is absent in chloroplasts. The complex with the internal signal sequence of LHCPs reveals that cpSRP43 specifically recognizes a DPLG peptide motif. We describe how cpSPR43 adapts the universally conserved SRP system to posttranslational targeting and insertion of the LHCP family of membrane proteins. 相似文献
10.
Jenner L Romby P Rees B Schulze-Briese C Springer M Ehresmann C Ehresmann B Moras D Yusupova G Yusupov M 《Science (New York, N.Y.)》2005,308(5718):120-123
The ribosome of Thermus thermophilus was cocrystallized with initiator transfer RNA (tRNA) and a structured messenger RNA (mRNA) carrying a translational operator. The path of the mRNA was defined at 5.5 angstroms resolution by comparing it with either the crystal structure of the same ribosomal complex lacking mRNA or with an unstructured mRNA. A precise ribosomal environment positions the operator stem-loop structure perpendicular to the surface of the ribosome on the platform of the 30S subunit. The binding of the operator and of the initiator tRNA occurs on the ribosome with an unoccupied tRNA exit site, which is expected for an initiation complex. The positioning of the regulatory domain of the operator relative to the ribosome elucidates the molecular mechanism by which the bound repressor switches off translation. Our data suggest a general way in which mRNA control elements must be placed on the ribosome to perform their regulatory task. 相似文献
11.
Schotte F Lim M Jackson TA Smirnov AV Soman J Olson JS Phillips GN Wulff M Anfinrud PA 《Science (New York, N.Y.)》2003,300(5627):1944-1947
We report picosecond time-resolved x-ray diffraction from the myoglobin (Mb) mutant in which Leu29 is replaced by Phe (L29Fmutant). The frame-by-frame structural evolution, resolved to 1.8 angstroms, allows one to literally "watch" the protein as it executes its function. Time-resolved mid-infrared spectroscopy of flash-photolyzed L29F MbCO revealed a short-lived CO intermediate whose 140-ps lifetime is shorter than that found in wild-type protein by a factor of 1000. The electron density maps of the protein unveil transient conformational changes far more dramatic than the structural differences between the carboxy and deoxy states and depict the correlated side-chain motion responsible for rapidly sweeping CO away from its primary docking site. 相似文献
12.
[目的]利用虚拟筛选技术研究粟米草中抑制TNF-α活性成分。[方法]采用计算机虚拟筛选手段,分子对接打分技术,研究粟米草中成分的三维结构同TNF-α受体的相互作用。[结果]以TNF-α为靶标与粟米草中分离到的化合物进行分子对接,从中筛选到4种活性较好的化合物,分别为quercetin、kaemferol、rutin、β-sitosterol。[结论]通过虚拟筛选技术寻找到粟米草中抑制TNF-α活性成分,为探索粟米草抗心律失常的有效成分提供理论和实验基础。 相似文献
13.
Rickert M Wang X Boulanger MJ Goriatcheva N Garcia KC 《Science (New York, N.Y.)》2005,308(5727):1477-1480
Interleukin-2 (IL-2) is an immunoregulatory cytokine that binds sequentially to the alpha (IL-2Ralpha), beta (IL-2Rbeta), and common gamma chain (gammac) receptor subunits. Here we present the 2.8 angstrom crystal structure of a complex between human IL-2 and IL-2Ralpha, which interact in a docking mode distinct from that of other cytokine receptor complexes. IL-2Ralpha is composed of strand-swapped "sushi-like" domains, unlike the classical cytokine receptor fold. As a result of this domain swap, IL-2Ralpha uses a composite surface to dock into a groove on IL-2 that also serves as a binding site for antagonist drugs. With this complex, we now have representative structures for each class of hematopoietic cytokine receptor-docking modules. 相似文献
14.
利用3种同源蛋白结构,通过同源模建的方法构建N-甲基-D-天冬氨酸(NMDA)受体NR2B亚基的三维构象,并对模建蛋白进行多种参数的验证.结果表明:在构建的模型中,conantokin-G(Con-G,NR2B亚基受体拮抗剂)与NR2B对接后,Con-G能很好地嵌入到NR2B亚基激动剂结合部位,并且Con-G的构象基本不变,仍保持α-螺旋构象;Con-G的E2、Gla4、L5、Q9、I12和Q13,NR2B的E420、S421、D423、K458和D715是参与相互作用的重要氨基酸,它们之间形成了一些重要的氢键.该模型的建立对预测NMDA受体与其配体的相互作用具有一定的作用,为设计新的NMDA受体激动剂和拮抗剂提供了重要线索. 相似文献
15.
16.
Using the atomic structures of the large ribosomal subunit from Haloarcula marismortui and its complexes with two substrate analogs, we establish that the ribosome is a ribozyme and address the catalytic properties of its all-RNA active site. Both substrate analogs are contacted exclusively by conserved ribosomal RNA (rRNA) residues from domain V of 23S rRNA; there are no protein side-chain atoms closer than about 18 angstroms to the peptide bond being synthesized. The mechanism of peptide bond synthesis appears to resemble the reverse of the acylation step in serine proteases, with the base of A2486 (A2451 in Escherichia coli) playing the same general base role as histidine-57 in chymotrypsin. The unusual pK(a) (where K(a) is the acid dissociation constant) required for A2486 to perform this function may derive in part from its hydrogen bonding to G2482 (G2447 in E. coli), which also interacts with a buried phosphate that could stabilize unusual tautomers of these two bases. The polypeptide exit tunnel is largely formed by RNA but has significant contributions from proteins L4, L22, and L39e, and its exit is encircled by proteins L19, L22, L23, L24, L29, and L31e. 相似文献
17.
Weixlbaumer A Jin H Neubauer C Voorhees RM Petry S Kelley AC Ramakrishnan V 《Science (New York, N.Y.)》2008,322(5903):953-956
The termination of protein synthesis occurs through the specific recognition of a stop codon in the A site of the ribosome by a release factor (RF), which then catalyzes the hydrolysis of the nascent protein chain from the P-site transfer RNA. Here we present, at a resolution of 3.5 angstroms, the crystal structure of RF2 in complex with its cognate UGA stop codon in the 70S ribosome. The structure provides insight into how RF2 specifically recognizes the stop codon; it also suggests a model for the role of a universally conserved GGQ motif in the catalysis of peptide release. 相似文献
18.
Dunkle JA Wang L Feldman MB Pulk A Chen VB Kapral GJ Noeske J Richardson JS Blanchard SC Cate JH 《Science (New York, N.Y.)》2011,332(6032):981-984
During protein synthesis, the ribosome controls the movement of tRNA and mRNA by means of large-scale structural rearrangements. We describe structures of the intact bacterial ribosome from Escherichia coli that reveal how the ribosome binds tRNA in two functionally distinct states, determined to a resolution of ~3.2 angstroms by means of x-ray crystallography. One state positions tRNA in the peptidyl-tRNA binding site. The second, a fully rotated state, is stabilized by ribosome recycling factor and binds tRNA in a highly bent conformation in a hybrid peptidyl/exit site. The structures help to explain how the ratchet-like motion of the two ribosomal subunits contributes to the mechanisms of translocation, termination, and ribosome recycling. 相似文献
19.
In bacteria, the hybrid transfer-messenger RNA (tmRNA) rescues ribosomes stalled on defective messenger RNAs (mRNAs). However, certain gram-negative bacteria have evolved proteins that are capable of rescuing stalled ribosomes in a tmRNA-independent manner. Here, we report a 3.2 angstrom-resolution crystal structure of the rescue factor YaeJ bound to the Thermus thermophilus 70S ribosome in complex with the initiator tRNA(i)(fMet) and a short mRNA. The structure reveals that the C-terminal tail of YaeJ functions as a sensor to discriminate between stalled and actively translating ribosomes by binding in the mRNA entry channel downstream of the A site between the head and shoulder of the 30S subunit. This allows the N-terminal globular domain to sample different conformations, so that its conserved GGQ motif is optimally positioned to catalyze the hydrolysis of peptidyl-tRNA. This structure gives insights into the mechanism of YaeJ function and provides a basis for understanding how it rescues stalled ribosomes. 相似文献
20.
Siridechadilok B Fraser CS Hall RJ Doudna JA Nogales E 《Science (New York, N.Y.)》2005,310(5753):1513-1515
Protein synthesis in mammalian cells requires initiation factor eIF3, a approximately 750-kilodalton complex that controls assembly of 40S ribosomal subunits on messenger RNAs (mRNAs) bearing either a 5'-cap or an internal ribosome entry site (IRES). Cryo-electron microscopy reconstructions show that eIF3, a five-lobed particle, interacts with the hepatitis C virus (HCV) IRES RNA and the 5'-cap binding complex eIF4F via the same domain. Detailed modeling of eIF3 and eIF4F onto the 40S ribosomal subunit reveals that eIF3 uses eIF4F or the HCV IRES in structurally similar ways to position the mRNA strand near the exit site of 40S, promoting initiation complex assembly. 相似文献