scholarly journals Interface size drives cotranslational assembly of protein complexes

2021 ◽  
Author(s):  
Mihaly Badonyi ◽  
Joseph A Marsh
Keyword(s):  
Structure Data ◽  
Protein Complexes ◽  
Ribosome Profiling ◽  
Specific Structure ◽  
Complex Assembly ◽  
Nascent Chain ◽  
Assembly Pathways ◽  
Biological Context

Assembly pathways of protein complexes should be precise and efficient to minimise misfolding and unwanted interactions with other proteins in the cell. One way to achieve this is by seeding complex assembly during translation via nascent chain engagement. Here, we considered the possibility that the propensity of subunits to cotranslationally assemble is ingrained within the interface hierarchy of protein complexes. Using a combination of proteome-specific structure data and assembly-onset positions determined by ribosome profiling, we show that larger interfaces are prioritised in the course of cotranslational assembly. We observe that this effect is not exclusive to homomeric complexes, but appears to drive the assembly of heteromeric subunits, to the extent that interface size differences are detectable between N and C-terminal locations, with the former being larger on average. We provide explanations to this phenomenon and discuss its importance in a biological context.

scholarly journals Co-translational assembly counteracts promiscuous interactions

2021 ◽  
Author(s):  
Maximilian Seidel ◽  
Anja Becker ◽  
Filipa Pereira ◽  
Jonathan J.M. Landry ◽  
Nayara Trevisan Doimo de Azevedo ◽  
...  
Keyword(s):  
Protein Complexes ◽  
Coiled Coils ◽  
Ribosome Profiling ◽  
Structural Motifs ◽  
Short Linear Motifs ◽  
Physiological Relevance ◽  
Nascent Chain ◽  
Regulatory Complexity ◽  
Linear Motifs ◽  
Assembly Pathways

During the co-translational assembly of protein complexes, a fully synthesized subunit engages with the nascent chain of a newly synthesized interaction partner. Such events are thought to contribute to productive assembly, but their exact physiological relevance remains underexplored. Here, we examined structural motifs contained in nucleoporins for their potential to facilitate co-translational assembly. We experimentally tested candidate structural motifs and identified several previously unknown co-translational interactions. We demonstrate by selective ribosome profiling that domain invasion motifs of beta-propellers, coiled-coils, and short linear motifs act as co-translational assembly domains. Such motifs are often contained in proteins that are members of multiple complexes (moonlighters) and engage with closely related paralogs. Surprisingly, moonlighters and paralogs assembled co-translationally in only one but not all of the relevant assembly pathways. Our results highlight the regulatory complexity of assembly pathways.

scholarly journals Ribosome elongation kinetics of consecutively charged residues are coupled to electrostatic force

2021 ◽  
Author(s):  
Sarah E Leininger ◽  
Judith Rodriguez ◽  
Quyen V Vu ◽  
Yang Jiang ◽  
Ma Suan Li ◽  
...  
Keyword(s):  
Amino Acid ◽  
Conformational Changes ◽  
Electrostatic Force ◽  
Ribosome Profiling ◽  
Peptide Bond Formation ◽  
Translation Speed ◽  
Charged Residues ◽  
Nascent Chain ◽  
Chain Conformations

The speed of protein synthesis can dramatically change when consecutively charged residues are incorporated into an elongating nascent protein by the ribosome. The molecular origins of this class of allosteric coupling remain unknown. We demonstrate, using multi-scale simulations, that positively charged residues generate large forces that pull the P-site amino acid away from the A-site amino acid. Negatively charged residues generate forces of similar magnitude but opposite direction. And that these conformational changes, respectively, raise and lower the transition state barrier height to peptide bond formation, explaining how charged residues mechanochemically alter translation speed. This mechanochemical mechanism is consistent with in vivo ribosome profiling data exhibiting a proportionality between translation speed and the number of charged residues, experimental data characterizing nascent chain conformations, and a previously published cryo-EM structure of a ribosome-nascent chain complex containing consecutive lysines. These results expand the role of mechanochemistry in translation, and provide a framework for interpreting experimental results on translation speed.

The landscape of translational stall sites in bacteria revealed by monosome and disome profiling

RNA ◽  
2021 ◽  
pp. rna.078188.120
Author(s):  
Tomoya Fujita ◽  
Takeshi Yokoyama ◽  
Mikako Shirouzu ◽  
Hideki Taguchi ◽  
Takuhiro Ito ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Low Frequency ◽  
Ribosome Profiling ◽  
Reporter Assay ◽  
Ribosome Stalling ◽  
Peptidyl Transfer ◽  
Cryo Electron Microscopy ◽  
Nascent Chain ◽  
Biochemical Validation ◽  
Trna Sequencing

Ribosome pauses are associated with various cotranslational events and determine the fate of mRNAs and proteins. Thus, the identification of precise pause sites across the transcriptome is desirable; however, the landscape of ribosome pauses in bacteria remains ambiguous. Here, we harness monosome and disome (or collided ribosome) profiling strategies to survey ribosome pause sites in Escherichia coli. Compared to eukaryotes, ribosome collisions in bacteria showed remarkable differences: a low frequency of disomes at stop codons, collisions occurring immediately after 70S assembly on start codons, and shorter queues of ribosomes trailing upstream. The pause sites corresponded with the biochemical validation by integrated nascent chain profiling (iNP) to detect polypeptidyl-tRNA, an elongation intermediate. Moreover, the subset of those sites showed puromycin resistance, presenting slow peptidyl transfer. Among the identified sites, the ribosome pause at Asn586 of ycbZ was validated by biochemical reporter assay, tRNA sequencing (tRNA-Seq), and cryo-electron microscopy (cryo-EM) experiments. Our results provide a useful resource for ribosome stalling sites in bacteria

scholarly journals Precursor MicroRNA-Programmed Silencing Complex Assembly Pathways in Mammals

2012 ◽  
Vol 46 (4) ◽  
pp. 507-517 ◽  
Author(s):  
Xuhang Liu ◽  
Dong-Yan Jin ◽  
Michael T. McManus ◽  
Zissimos Mourelatos
Keyword(s):  
Complex Assembly ◽  
Assembly Pathways

scholarly journals Integrated in vivo and in vitro nascent chain profiling reveals widespread translational pausing

2016 ◽  
Vol 113 (7) ◽  
pp. E829-E838 ◽  
Author(s):  
Yuhei Chadani ◽  
Tatsuya Niwa ◽  
Shinobu Chiba ◽  
Hideki Taguchi ◽  
Koreaki Ito
Keyword(s):  
Escherichia Coli ◽  
Membrane Proteins ◽  
Ribosome Profiling ◽  
Cytosolic Proteins ◽  
Nascent Polypeptide ◽  
Nascent Chain ◽  
Chemical Trait

Although the importance of the nonuniform progression of elongation in translation is well recognized, there have been few attempts to explore this process by directly profiling nascent polypeptides, the relevant intermediates of translation. Such approaches will be essential to complement other approaches, including ribosome profiling, which is extremely powerful but indirect with respect to the actual translation processes. Here, we use the nascent polypeptide's chemical trait of having a covalently attached tRNA moiety to detect translation intermediates. In a case study,Escherichia coliSecA was shown to undergo nascent polypeptide-dependent translational pauses. We then carried out integrated in vivo and in vitro nascent chain profiling (iNP) to characterize 1,038 proteome members ofE.colithat were encoded by the first quarter of the chromosome with respect to their propensities to accumulate polypeptidyl–tRNA intermediates. A majority of them indeed undergo single or multiple pauses, some occurring only in vitro, some occurring only in vivo, and some occurring both in vivo and in vitro. Thus, translational pausing can be intrinsically robust, subject to in vivo alleviation, or require in vivo reinforcement. Cytosolic and membrane proteins tend to experience different classes of pauses; membrane proteins often pause multiple times in vivo. We also note that the solubility of cytosolic proteins correlates with certain categories of pausing. Translational pausing is widespread and diverse in nature.

scholarly journals Evidence for Membrane Complex Assembly in Nanoelectrospray Generated Lipid Bilayers

2019 ◽  
Author(s):  
Matthias Wilm
Keyword(s):  
Membrane Proteins ◽  
Membrane Protein ◽  
Lipid Bilayers ◽  
Layered Structure ◽  
Protein Complexes ◽  
Complex Assembly ◽  
Membrane Complex ◽  
Detergent Extraction ◽  
Membrane Protein Complexes

1.AbstractNanoelectrospray can be used to generate a layered structure consisting of bipolar lipids, detergent-solubilized membrane proteins, and glycerol that self-assembles upon detergent extraction into one extended layer of a protein containing membrane. This manuscript presents the first evidence that this method might allow membrane protein complexes to assemble in this process.

scholarly journals Emerging role of long noncoding RNA-encoded micropeptides in cancer

2020 ◽  
Vol 20 (1) ◽  
Author(s):  
Mujie Ye ◽  
Jingjing Zhang ◽  
Meng Wei ◽  
Baihui Liu ◽  
Kuiran Dong
Keyword(s):  
Noncoding Rna ◽  
Cancer Metabolism ◽  
Chain Complex ◽  
Ribosome Profiling ◽  
Open Reading Frames ◽  
Clinical Value ◽  
Anti Cancer ◽  
Nascent Chain ◽  
Reading Frames

Abstract Increasing evidence has indicated that long noncoding RNAs (lncRNAs) play various important roles in the development of cancers. The widespread applications of ribosome profiling and ribosome nascent chain complex sequencing revealed that some short open reading frames of lncRNAs have micropeptide-coding potential. The resulting micropeptides have been shown to participate in N6-methyladenosine modification, tumor angiogenesis, cancer metabolism, and signal transduction. This review summarizes current information regarding the reported roles of lncRNA-encoded micropeptides in cancer, and explores the potential clinical value of these micropeptides in the development of anti-cancer drugs and prognostic tumor biomarkers.

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