A Novel Protein Domain Induces High Affinity Selenocysteine Insertion Sequence Binding and Elongation Factor Recruitment

The IL-2 toxin-mediated inhibition of protein synthesis in high affinity IL-2-R-positive murine and human T cell lines has been examined. Both excess free IL-2 and mAb to the Tac epitope of the p55 subunit of IL-2-R are shown to block the action of IL-2 toxin; whereas, agents that interact with other receptors or antigens on the T cell surface have no effect. We show that IL-2 toxin, like diphtheria toxin, must pass through an acidic vesicle in order to intoxicate target T cells. Finally, we demonstrate that the IL-2 toxin-mediated inhibition of protein synthesis in both human and murine T cells that bear the high affinity IL-2-R is due to the classic diphtheria toxin fragment A-catalyzed ADP ribosylation of elongation factor 2.

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Synthesis runs counter to directional folding of a nascent protein domain

Nature Communications ◽

10.1038/s41467-020-18921-8 ◽

2020 ◽

Vol 11 (1) ◽

Author(s):

Xiuqi Chen ◽

Nandakumar Rajasekaran ◽

Kaixian Liu ◽

Christian M. Kaiser

Keyword(s):

Single Molecule ◽

Molten Globule ◽

Elongation Factor ◽

Protein Domain ◽

Folding Pathway ◽

Native Structure ◽

Folding Intermediates ◽

C Terminus

Abstract Folding of individual domains in large proteins during translation helps to avoid otherwise prevalent inter-domain misfolding. How folding intermediates observed in vitro for the majority of proteins relate to co-translational folding remains unclear. Combining in vivo and single-molecule experiments, we followed the co-translational folding of the G-domain, encompassing the first 293 amino acids of elongation factor G. Surprisingly, the domain remains unfolded until it is fully synthesized, without collapsing into molten globule-like states or forming stable intermediates. Upon fully emerging from the ribosome, the G-domain transitions to its stable native structure via folding intermediates. Our results suggest a strictly sequential folding pathway initiating from the C-terminus. Folding and synthesis thus proceed in opposite directions. The folding mechanism is likely imposed by the final structure and might have evolved to ensure efficient, timely folding of a highly abundant and essential protein.

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Synthesis runs counter to directional folding of a nascent protein domain

10.1101/2020.04.29.068593 ◽

2020 ◽

Cited By ~ 1

Author(s):

Xiuqi Chen ◽

Nandakumar Rajasekaran ◽

Kaixian Liu ◽

Christian M. Kaiser

Keyword(s):

Single Molecule ◽

Molten Globule ◽

Elongation Factor ◽

Protein Domain ◽

Folding Pathway ◽

Native Structure ◽

Folding Intermediates ◽

C Terminus

AbstractFolding of individual domains in large proteins during translation helps to avoid otherwise prevalent inter-domain misfolding. How folding intermediates observed in vitro for the majority of proteins relate to co-translational folding remains unclear. Combining in vivo and single-molecule experiments, we followed the co-translational folding of the G-domain, encompassing the first 293 amino acids of elongation factor G. Surprisingly, the domain remains unfolded until it is fully synthesized, without collapsing into molten globule-like states or forming stable intermediates. Upon fully emerging from the ribosome, the G-domain transitions to its stable native structure via folding intermediates. Our results suggest a strictly sequential folding pathway initiating from the C-terminus. Folding and synthesis thus proceed in opposite directions. The folding mechanism is likely imposed by the final structure and might have evolved to ensure efficient, timely folding of a highly abundant and essential protein.

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DWNN, a novel protein domain with a possible role in apoptosis and CTL-killing

Biochemical Society Transactions ◽

10.1042/bst028a230a ◽

2000 ◽

Vol 28 (5) ◽

pp. A230-A230

Author(s):

D. J. Rees ◽

D. J. Pugh

Keyword(s):

Protein Domain ◽

Novel Protein

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Recognition of the mRNA selenocysteine insertion sequence by the specialized translational elongation factor SELB.

Genes & Development ◽

10.1101/gad.8.3.376 ◽

1994 ◽

Vol 8 (3) ◽

pp. 376-385 ◽

Cited By ~ 27

Author(s):

S Ringquist ◽

D Schneider ◽

T Gibson ◽

C Baron ◽

A Bock ◽

...

Keyword(s):

Insertion Sequence ◽

Elongation Factor

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Antibody affinity versus dengue morphology influences neutralization

PLoS Pathogens ◽

10.1371/journal.ppat.1009331 ◽

2021 ◽

Vol 17 (2) ◽

pp. e1009331

Author(s):

Guntur Fibriansah ◽

Elisa X. Y. Lim ◽

Jan K. Marzinek ◽

Thiam-Seng Ng ◽

Joanne L. Tan ◽

...

Keyword(s):

Human Monoclonal Antibody ◽

Protein Domain ◽

E Proteins ◽

Virus Surface ◽

Exchange Mass Spectrometry ◽

High Affinity ◽

Hydrogen Deuterium ◽

Different Strains ◽

Dynamics Simulations ◽

Surface Morphologies

Different strains within a dengue serotype (DENV1-4) can have smooth, or “bumpy” surface morphologies with different antigenic characteristics at average body temperature (37°C). We determined the neutralizing properties of a serotype cross-reactive human monoclonal antibody (HMAb) 1C19 for strains with differing morphologies within the DENV1 and DENV2 serotypes. We mapped the 1C19 epitope to E protein domain II by hydrogen deuterium exchange mass spectrometry, cryoEM and molecular dynamics simulations, revealing that this epitope is likely partially hidden on the virus surface. We showed the antibody has high affinity for binding to recombinant DENV1 E proteins compared to those of DENV2, consistent with its strong neutralizing activities for all DENV1 strains tested regardless of their morphologies. This finding suggests that the antibody could out-compete E-to-E interaction for binding to its epitope. In contrast, for DENV2, HMAb 1C19 can only neutralize when the epitope becomes exposed on the bumpy-surfaced particle. Although HMAb 1C19 is not a suitable therapeutic candidate, this study with HMAb 1C19 shows the importance of choosing a high-affinity antibody that could neutralize diverse dengue virus morphologies for therapeutic purposes.

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Synthesis of a Novel Bicyclic Scaffold Inspired by the Antifungal Natural Product Sordarin

10.26434/chemrxiv.6394409.v1 ◽

2018 ◽

Author(s):

Yibiao Wu ◽

Chris Dockendorff

Keyword(s):

Carboxylic Acid ◽

Natural Product ◽

Elongation Factor ◽

Alder Reaction ◽

Diels Alder ◽

High Affinity ◽

Diels Alder Reaction ◽

Elongation Factor 2 ◽

Eukaryotic Elongation Factor 2 ◽

Eukaryotic Elongation Factor

A simplified bicyclic scaffold inspired by the antifungal natural product sordarin was designed and synthesized which maintains the carboxylic acid/aldehyde (or nitrile) pharmacophore. A densely functionalized chiral cyclopentadiene was constructed in 8 steps and utilized in a Diels-Alder reaction with acrylonitrile. The resulting [2.2.1]cycloheptene was transformed into a scaffold possessing vicinal carboxylic acid and nitrile groups, with orientations predicted to provide high affinity for the fungal protein eukaryotic elongation factor 2 (eEF2).

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Bioinformatic Prediction of an tRNASec Gene Nested inside an Elongation Factor SelB Gene in Alphaproteobacteria

International Journal of Molecular Sciences ◽

10.3390/ijms22094605 ◽

2021 ◽

Vol 22 (9) ◽

pp. 4605

Author(s):

Takahito Mukai

Keyword(s):

Insertion Sequence ◽

Trna Gene ◽

Stop Codon ◽

Elongation Factor ◽

Selenium Deficiency ◽

Open Reading Frame ◽

Start Codon ◽

Sequence Element ◽

Reading Frame ◽

Insertion Sequence Element

In bacteria, selenocysteine (Sec) is incorporated into proteins via the recoding of a particular codon, the UGA stop codon in most cases. Sec-tRNASec is delivered to the ribosome by the Sec-dedicated elongation factor SelB that also recognizes a Sec-insertion sequence element following the codon on the mRNA. Since the excess of SelB may lead to sequestration of Sec-tRNASec under selenium deficiency or oxidative stress, the expression levels of SelB and tRNASec should be regulated. In this bioinformatic study, I analyzed the Rhizobiales SelB species because they were annotated to have a non-canonical C-terminal extension. I found that the open reading frame (ORF) of diverse Alphaproteobacteria selB genes includes an entire tRNASec sequence (selC) and overlaps with the start codon of the downstream ORF. A remnant tRNASec sequence was found in the Sinorhizobium melilotiselB genes whose products have a shorter C-terminal extension. Similar overlapping traits were found in Gammaproteobacteria and Nitrospirae. I hypothesized that once the tRNASec moiety is folded and processed, the expression of the full-length SelB may be repressed. This is the first report on a nested tRNA gene inside a protein ORF in bacteria.

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