Protein design

  • Article
    | Open Access

    Major Histocompatibility Complex (MHC) class I molecules present tightly binding peptides on the cell surface for recognition by cytotoxic T cells. Here, the authors present the crystal structures of a disulfide-stabilized human MHC class I molecule in the peptide-free state and bound with dipeptides, and find that peptide binding is accompanied by concerted conformational switches of the amino acid side chains in the binding pockets.

    • Raghavendra Anjanappa
    • , Maria Garcia-Alai
    •  & Rob Meijers
  • Article
    | Open Access

    CRISPR homing gene drives are highly invasive and can fail due to the rapid evolution of resistance. Here the authors present TARE drive, inspired by naturally occurring selfish genetic elements, which is less vulnerable to resistance and can potentially be confined to a target population.

    • Jackson Champer
    • , Esther Lee
    •  & Philipp W. Messer
  • Article
    | Open Access

    Many reagents have been developed for cysteine-specific protein modification. However, few of them allow for multi-functionalization of a single Cys residue and disulfide bridging bioconjugation. Here the authors report 3-bromo-5-methylene pyrrolones as a simple, robust and versatile class of reagents for cysteine-specific protein modification.

    • Yingqian Zhang
    • , Chuanlong Zang
    •  & Chuanzheng Zhou
  • Article
    | Open Access

    Protein tyrosine phosphatases regulate many cellular processes but are difficult to study in their native context. Here the authors develop an approach for using light to control the activity of a disease-relevant phosphatase without interfering with its native cellular organization.

    • Akarawin Hongdusit
    • , Peter H. Zwart
    •  & Jerome M. Fox
  • Article
    | Open Access

    Antibodies expressed in the cytosol often form insoluble aggregates, which makes it hard to target intracellular proteins. Here the authors engineer an ultra-stable cytoplasmic antibody (STAND) with a low isoelectric point that can be used in vivo.

    • Hiroyuki Kabayama
    • , Makoto Takeuchi
    •  & Katsuhiko Mikoshiba
  • Article
    | Open Access

    Monomeric near-infrared (NIR) fluorescent proteins (FPs) from bacterial phytochromes bring potential advantages, but their brightness in cells is lower than dimeric NIR FPs. Here the authors develop enhanced monomeric NIR FPs enabling imaging across different scales without the trade-off between brightness and monomeric state.

    • Mikhail E. Matlashov
    • , Daria M. Shcherbakova
    •  & Vladislav V. Verkhusha
  • Article
    | Open Access

    Multiple sequence alignments of proteins carry information about evolution, the protein’s fitness landscape and its stability in the face of mutations. Here, the authors demonstrate the utility of latent space models learned using variational autoencoders to infer these properties from sequences.

    • Xinqiang Ding
    • , Zhengting Zou
    •  & Charles L. Brooks III
  • Article
    | Open Access

    Lee et al. report an engineered IgG1 Fc domain that behaves like an hFcRn binding pH toggle switch. The authors show that this new half-life extension Fc domain confers improved pharmacokinetics in new humanized knock-in mouse strains that recapitulate the key processes for antibody persistence in circulation.

    • Chang-Han Lee
    • , Tae Hyun Kang
    •  & George Georgiou
  • Article
    | Open Access

    Developing inhibitors that target specific protein-protein interactions (PPIs) is challenging. Here, the authors show that target selectivity and PPI blocking can be achieved simultaneously with PPI inhibitors that contain two functional modules, and create a paralog-selective PSD-95 inhibitor as proof-of-concept.

    • Charlotte Rimbault
    • , Kashyap Maruthi
    •  & Matthieu Sainlos
  • Article
    | Open Access

    Increasing TCR cell surface expression can potentiate T cell responses to low-concentrations of antigen. Here the authors identify aminoacids in human TCR variable domains that impact its surface expression, and demonstrate how editing these residues can improve T cell activation and effector function without altering antigen specificity.

    • Sharyn Thomas
    • , Fiyaz Mohammed
    •  & Hans J. Stauss
  • Article
    | Open Access

    Conserved regions of the antibody molecule impact its downstream biological effects. Here the authors show that a rigid hinge conformation increases the agonistic activity of CD40 and DR5 antibodies, distinctly from FcγR-binding, suggesting that the hinge and FcR binding regions can be separately modified to optimize therapies.

    • Xiaobo Liu
    • , Yingjie Zhao
    •  & Fubin Li
  • Article
    | Open Access

    Nanoparticles are a promising approach to increase immunogenicity of protein antigens for vaccines. Here, Brouwer et al. design self-assembling, two-component protein NPs that present native-like SOSIP trimers of HIV envelope protein and determine immunogenicity in a small animal model.

    • Philip J. M. Brouwer
    • , Aleksandar Antanasijevic
    •  & Rogier W. Sanders
  • Article
    | Open Access

    Proteins have been used in the synthesis of magnetic nanoparticles but issues with aggregation limit this application. Here, the authors report on the synthesis of coiled proteins that display the active loop of the natural proteins to avoid aggregation and investigate the application in nanoparticle synthesis.

    • Andrea E. Rawlings
    • , Lori A. Somner
    •  & Sarah S. Staniland
  • Article
    | Open Access

    Polyglutamine (polyQ) tracts are low-complexity regions and their expansion is linked to certain neurodegenerative diseases. Here the authors combine experimental and computational approaches to find that the length of the androgen receptor polyQ tract correlates with its helicity and show that the polyQ helical structure is stabilized by hydrogen bonds between the Gln side chains and main chain carbonyl groups.

    • Albert Escobedo
    • , Busra Topal
    •  & Xavier Salvatella
  • Article
    | Open Access

    Proteins smaller than about 50 kDa are currently too small to be imaged at high resolution by cryo‐electron microscopy (cryo‐EM). Here authors design a protein scaffold that binds 12 copies of a small 26 kDa protein (GFP), which allowed visualizing GFP at a resolution of 3.8Å by cryo‐EM.

    • Yuxi Liu
    • , Duc T. Huynh
    •  & Todd O. Yeates
  • Article
    | Open Access

    Genome editing often requires cleavage within a narrow sequence window. Here the authors develop an expanded set of zinc finger nuclease architectures that increase the available configurations by a factor of 64 and can target almost every base at loci of therapeutic significance.

    • David E. Paschon
    • , Stephanie Lussier
    •  & Edward J. Rebar
  • Article
    | Open Access

    Optical control over receptor tyrosine kinase (RTK) activity can find application in both basic biology and synthetic biology. Here the authors combine the photosensory module of DrBphP bacterial phytochrome with the Trk family of RTKs to obtain Trks that are reversibly switchable with near-infrared and far-red light.

    • Anna V. Leopold
    • , Konstantin G. Chernov
    •  & Vladislav V. Verkhusha
  • Article
    | Open Access

    Metabolic engineering usually focuses on manipulating enzyme(s) within a single pathway. Here, the authors show that a promiscuous enzymatic activity-based multiple-pathway design can minimize cell metabolic burden and allow carbon flow rearrangement, leading to efficient hydroxytyrosol biosynthesis.

    • Wei Chen
    • , Jun Yao
    •  & Shuang-Yan Tang
  • Article
    | Open Access

    The Cas12b family of CRISPR nucleases has been underutilized in mammalian cells due to the high temperature requirement of known members. Here the authors engineer BhCas12b to overcome this limitation for robust and specific genome editing applications in human cells.

    • Jonathan Strecker
    • , Sara Jones
    •  & Feng Zhang
  • Article
    | Open Access

    Near-infrared (NIR) fluorescent proteins (FPs) offer advantages for spectral multiplexing and deep-tissue imaging. Here the authors engineer a smaller NIR FP based on the unexplored cyanobacteriochrome photoreceptor and demonstrate its use in various applications in cell culture as well as whole-body imaging in vivo in mice.

    • Olena S. Oliinyk
    • , Anton A. Shemetov
    •  & Vladislav V. Verkhusha
  • Article
    | Open Access

    The use of enzymes as drugs requires that they persist within target tissues over therapeutically relevant time frames. Here the authors use a galectin-3 fusion to anchor enzymes at injection sites for up to 14 days.

    • Shaheen A. Farhadi
    • , Evelyn Bracho-Sanchez
    •  & Gregory A. Hudalla
  • Article
    | Open Access

    Mutating natural enzymes is effective in broadening the substrate or product range, but generally leads to reduced titers. Here the authors engineer hybrid polyketide synthases for efficient production of short-chain ketones from plant biomass hydrolysates in Streptomyces, which can increase the octane of gasoline.

    • Satoshi Yuzawa
    • , Mona Mirsiaghi
    •  & Jay D. Keasling
  • Article
    | Open Access

    Methods to directly label active neurons are still lacking. Here the authors develop CaMPARI2, a photoconvertible fluorescent protein sensor for neuronal activity with improved brightness and calcium binding kinetics, as well as an antibody to amplify the activated sensor signal in fixed samples.

    • Benjamien Moeyaert
    • , Graham Holt
    •  & Eric R. Schreiter
  • Article
    | Open Access

    Higher order coiled coils with five or more helices can form α-helical barrels. Here the authors show that placing β-branched aliphatic residues along the lumen yields stable and open α-helical barrels, which is of interest for the rational design of functional proteins; whereas, the absence of β-branched side chains leads to unusual low-symmetry α-helical bundles.

    • Guto G. Rhys
    • , Christopher W. Wood
    •  & Derek N. Woolfson
  • Article
    | Open Access

    Designing split protein approaches is time consuming and often results in high background activity due to spontaneous assembly. Here the authors present an automated approach which uses a split energy scoring function to identify optimal protein split sites and reduces spontaneous assembly.

    • Onur Dagliyan
    • , Andrey Krokhotin
    •  & Nikolay V. Dokholyan
  • Article
    | Open Access

    Characterizing the binding selectivity landscape of interacting proteins is crucial in protein engineering. Here the authors use multi-target selective library screening and in silico next-generation sequencing to map the binding landscape of proteins and produce improved proteases inhibitors.

    • Si Naftaly
    • , Itay Cohen
    •  & Niv Papo
  • Article
    | Open Access

    Previous efforts to assemble Rubisco within a cyanobacterial carboxysome-derived protein shell in plant chloroplasts to concentrate CO2 have been unsuccessful. Here, Long et al. produce carboxysomes in tobacco chloroplasts that encapsulate the introduced Rubisco and enable autotrophic growth at elevated CO2.

    • Benedict M. Long
    • , Wei Yih Hee
    •  & G. Dean Price
  • Article
    | Open Access

    Bacterial microcompartments (BMCs) are protein-bound organelles encapsulating segments of metabolic pathways. Here the authors utilize specific de novo coiled-coil protein-protein interactions to display proteins on the outer or inner surface of BMCs.

    • Matthew J. Lee
    • , Judith Mantell
    •  & Martin J. Warren
  • Article
    | Open Access

    Current approaches to conditionally deplete target proteins require site-specific genetic engineering or have poor temporal control. Here the authors overcome these limitations by combining the AID system with nanobodies to reversibly degrade GFP-tagged proteins in living cells and zebrafish.

    • Katrin Daniel
    • , Jaroslav Icha
    •  & Jörg Mansfeld
  • Article
    | Open Access

    His-tagged proteins can undergo N-terminal acylation as an undesired side-reaction. Here, the authors utilize this to develop a method for highly selective acylation and further modification of peptides and proteins using an optimized His sequence and 4-methoxyphenyl esters as acyl donors.

    • Manuel C. Martos-Maldonado
    • , Christian T. Hjuler
    •  & Knud J. Jensen
  • Article
    | Open Access

    Computationally designed enzymes often show lower activity or stability than their natural counterparts. Here, the authors present an evolution-inspired method for automated enzyme design, creating stable enzymes with accurate active site architectures and wild-type-like activities.

    • Gideon Lapidoth
    • , Olga Khersonsky
    •  & Sarel J. Fleishman
  • Article
    | Open Access

    Pentatricopeptide repeat proteins bind single-stranded RNA and have been used to study ssRNA biology. Here the authors co-opt these proteins to target ssDNA and demonstrate specific binding of telomere sequences, the structural basis for ssDNA wrapping, and use them as potent telomerase inhibitors.

    • Henrik Spåhr
    • , Tiongsun Chia
    •  & Oliver Rackham
  • Article
    | Open Access

    CXCR2 antagonism has been shown to be anti-arthritic, but anti-chemokine therapies usually fail in the clinic owing to redundancy in chemokine-receptor interactions. Here the authors develop single-chain antibodies with multiple chemokine specificities to achieve high affinity and broad specificity to mouse and human CXC chemokines with efficacy in a K/BxN serum transfer model of arthritis.

    • Alessandro Angelini
    • , Yoshishige Miyabe
    •  & K. Dane Wittrup
  • Article
    | Open Access

    Cellobiohydrolases (CBHs) are critical for natural and industrial biomass degradation but their structure–activity relationships are not fully understood. Here, the authors present the biochemical and structural characterization of two CBHs, identifying protein regions that confer enhanced CBH activity.

    • Larry E. Taylor II
    • , Brandon C. Knott
    •  & Gregg T. Beckham
  • Article
    | Open Access

    Optimizing an enzyme usually requires testing thousands of variants, thus consuming large amounts of material and time. Here, the authors present a method that allows for measuring two different activities of the same enzyme simultaneously instead of doing two consecutive rounds of screening.

    • Fuqiang Ma
    • , Meng Ting Chung
    •  & Guang-Yu Yang
  • Article
    | Open Access

    With the growing threat of antibiotic resistance, unconventional approaches to antimicrobial discovery are needed. Here, the authors present a peptide topology that mimics virus architecture and assembles into antimicrobial capsids that disrupt bacterial membranes upon contact.

    • Emiliana De Santis
    • , Hasan Alkassem
    •  & Maxim G. Ryadnov
  • Article
    | Open Access

    Pathologies related to protein:protein interaction are hard to treat but cystine-dense peptides have the potential to disrupt such interactions. Here the authors develop a high-diversity mammalian cell screen for cystine-dense peptides with drug potential and use it to identify a YAP:TEAD inhibitor.

    • Zachary R. Crook
    • , Gregory P. Sevilla
    •  & James M. Olson
  • Article
    | Open Access

    E3 ubiquitin ligases regulate biological functions by ubiquitinating defined substrate proteins but overlapping specificities complicate the identification of E3-substrate relationships. Here, the authors construct an orthogonal UB transfer cascade and identify specific substrates of the E3 enzyme E6AP.

    • Yiyang Wang
    • , Xianpeng Liu
    •  & Jun Yin