Biochemistry

A broadly neutralizing biparatopic Nanobody protects mice from lethal challenge with SARS-CoV-2 variants of concern

1. Teresa R. Wagner
2. Daniel Schnepf
3. Julius Beer
4. Karin Klingel
5. Natalia Ruetalo
6. Philipp D. Kaiser
7. Daniel Junker
8. Martina Sauter
9. Bjoern Traenkle
10. Desiree I. Frecot
11. Matthias Becker
12. Nicole Schneiderhan-Marra
13. Annette Ohnemus
14. Martin Schwemmle
15. Michael Schindler
16. Ulrich Rothbauer

Reviewed by ScreenIT

Peptide Scanning of SARS-CoV and SARS-CoV-2 Spike Protein Subunit 1 Reveals Potential Additional Receptor Binding Sites

1. Weilin Lin
2. Jannatul Rafeya
3. Vanessa Roschewitz
4. David Smith
5. Adrian Keller
6. Yixin Zhang

Reviewed by ScreenIT

Framework for rapid comparison of extracellular vesicle isolation methods

1. Dmitry Ter-Ovanesyan
2. Maia Norman
3. Roey Lazarovits
4. Wendy Trieu
5. Ju-Hyun Lee
6. George M Church
7. David R Walt

• Curated by eLife

  Evaluation Summary:

  This manuscript describes a framework for rapidly quantifying relative extracellular vesicle (EV) yield and purity across isolation methods, with a focus on using size exclusion chromatography (SEC) for EV isolation from small volumes of pooled plasma and cerebrospinal fluid (CSF) samples. The authors used single molecule array (Simoa) assays for the quantification of EVs using three tetraspanins (CD9, CD63, and CD81), and report the outcomes of assessing EV yields and purity with respect to albumin by various SEC parameters (Sepharose size, column length, fractions collected). This is the first demonstration of the use of Simoa with three commonly used tetraspanins to measure EVs from small volumes of CSF, of great relevance to human CSF biomarker studies, but these methods could also be applied to compare EV isolation methods from other fluids such as cell culture media.

  (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 and Reviewer #3 agreed to share their names with the authors.)

Reviewed by eLife

Cryo-EM structure determination of small proteins by nanobody-binding scaffolds (Legobodies)

1. Xudong Wu
2. Tom A. Rapoport

Reviewed by ScreenIT

Native proline-rich motifs exploit sequence context to target actin-remodeling Ena/VASP protein ENAH

1. Theresa Hwang
2. Sara S Parker
3. Samantha M Hill
4. Robert A Grant
5. Meucci W Ilunga
6. Venkatesh Sivaraman
7. Ghassan Mouneimne
8. Amy E Keating

• Curated by eLife

  Evaluation Summary:

  The manuscript introduces a new molecular screen, MassTitr, to screen for long (36-mer) peptides derived from the human proteome that can bind a specific target. The method is demonstrated using the EVH1 domain of the actin-associated ENAH protein as target. About 100 peptides were isolated, and further analysis identified sequence features that contribute to the binding of the EVH1 domain by these peptides. The human proteome contains many short linear motifs of 4-6 residues that are critical for protein-protein interactions. The work here helps to better understand how the sequence surrounding such motifs contributes to protein-protein interactions.

  (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 and Reviewer #2 agreed to share their names with the authors.)

Reviewed by eLife

A distributed residue network permits conformational binding specificity in a conserved family of actin remodelers

1. Theresa Hwang
2. Sara S Parker
3. Samantha M Hill
4. Meucci W Ilunga
5. Robert A Grant
6. Ghassan Mouneimne
7. Amy E Keating

• Curated by eLife

  Evaluation Summary:

  This manuscript describes follow-up studies on a hit from a proteome-wide screen for peptides that can bind to the EVH1 domain of the ENAH protein, one of three highly similar Ena/VASP actin regulators. The hit investigated is from a protein called PCARE, which selectively binds to ENAH but not the other two members of the Ena/VASP family, EVL and VASP. The authors provide a good explanation for how this selectivity is achieved and develop a peptide, PCARE-Dual, that specifically binds ENAH more tightly, setting out the stage for developing potent and selective inhibitors of ENAH activity.

  (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 and Reviewer #2 agreed to share their name with the authors.)

Reviewed by eLife

Characterization of the ABC methionine transporter from Neisseria meningitidis reveals that lipidated MetQ is required for interaction

1. Naima G Sharaf
2. Mona Shahgholi
3. Esther Kim
4. Jeffrey Y Lai
5. David G VanderVelde
6. Allen T Lee
7. Douglas C Rees

• Curated by eLife

  Evaluation Summary:

  Sharaf and colleagues present an elegant structural and functional analysis of the Neisseria meningitidis ABC transporters MetQ/MetNI illustrating that the substrate binding protein MetQ requires N-terminal lipidation and a substrate (e.g. L-Met and other Met analogs) to stimulate the ATPase, presumably in order to transport the substrate across the inner membrane. This paper will be of broad interest to microbiologists and membrane physiologists who study periplasmic substrate binding proteins and transporter interactions in bacteria.

  (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #3 agreed to share their name with the authors.)

Reviewed by eLife

Rad53 checkpoint kinase regulation of DNA replication fork rate via Mrc1 phosphorylation

1. Allison W McClure
2. John FX Diffley

• Curated by eLife

  Evaluation Summary:

  This manuscript will be of interest to those interested in the regulation of DNA replication and those interested in how DNA damage impacts DNA replication. DNA replication must occur accurately to maintain genome integrity and also must be able to deal with DNA damage or metabolic conditions that induce replication fork stalling. Two key proteins involved in signaling such replication stress are Mrc1 and Rad53 kinase, and the authors use a powerful in vitro reconstitution system to make findings pertaining to these two proteins that are then supported by genetics.

  (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 and Reviewer #3 agreed to share their names with the authors.)

Reviewed by eLife

Small-molecule ligands can inhibit −1 programmed ribosomal frameshifting in a broad spectrum of coronaviruses

1. Sneha Munshi
2. Krishna Neupane
3. Sandaru M. Ileperuma
4. Matthew T.J. Halma
5. Jamie A. Kelly
6. Clarissa F. Halpern
7. Jonathan D. Dinman
8. Sarah Loerch
9. Michael T. Woodside

Reviewed by Rapid Reviews Infectious Diseases, ScreenIT

Witnessing the structural evolution of an RNA enzyme

1. Xavier Portillo
2. Yu-Ting Huang
3. Ronald R Breaker
4. David P Horning
5. Gerald F Joyce

• Curated by eLife

  Evaluation Summary:

  The plasticity of RNA folds and their ability to response to changes in selective pressure is a key aspect of understanding the evolution of life on this planet. The Class I ligase is a remarkably fast RNA ligase ribozyme that has been harnessed by a number of laboratories to power RNA polymerization. Thought by many to be the immutable catalytic core required for polymerization, Portillo et al. demonstrate evolutionary trajectories that result in a new and catalytically enhanced ligase core. An accumulation of mutations results in a the formation of a new pseudoknot structure immediately outside the active site of the ligase core. This new structure appears to more optimally position the P7-P6-P3 coaxially stacked stems of the ligase core with respect to the primer template substrate. Tracking the emergence of this new fold, which is correlated with an enhancement in RNA polymerization activity, is novel and interesting.

  (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. Reviewer #1 and Reviewer #3 agreed to share their names with the authors.)

Reviewed by eLife