AvrSr27 is a zinc-bound effector with a modular structure important for immune recognition.

Effector proteins are central to the success of plant pathogens, while immunity in host plants is driven by receptor-mediated recognition of these effectors. Understanding the molecular details of effector-receptor interactions is key for the engineering of novel immune receptors. Here, we experimentally determined the crystal structure of the Puccinia graminis f. sp. tritici (Pgt) effector AvrSr27, which was not accurately predicted using AlphaFold2. We characterised the role of the conserved cysteine residues in AvrSr27 using in vitro biochemical assays and examined Sr27-mediated recognition using transient expression in Nicotiana spp. and wheat protoplasts. The AvrSr27 structure contains a novel ß-strand rich modular fold consisting of two structurally similar domains that bind to Zn2+ ions. The N-terminal domain of AvrSr27 is sufficient for interaction with Sr27 and triggering cell death. We identified two Pgt proteins structurally related to AvrSr27 but with low sequence identity that can also associate with Sr27, albeit more weakly. Though only the full-length proteins, trigger Sr27-dependent cell death in transient expression systems. Collectively, our findings have important implications for utilising protein prediction platforms for effector proteins, and those embarking on bespoke engineering of immunity receptors as solutions to plant disease.

How Pore Architecture Regulates the Function of Nanoscale Protein Compartments.

Self-assembling proteins can form porous compartments that adopt well-defined architectures at the nanoscale. In nature, protein compartments act as semipermeable barriers to enable spatial separation and organization of complex biochemical processes. The compartment pores play a key role in their overall function by selectively controlling the influx and efflux of important biomolecular species. By engineering the pores, the functionality of compartments can be tuned to facilitate non-native applications, such as artificial nanoreactors for catalysis. In this review, we analyze how protein structure determines the porosity and impacts the function of both native and engineered compartments, highlighting the wealth of structural data recently obtained by cryo-EM and X-ray crystallography. Through this analysis, we offer perspectives on how current structural insights can inform future studies into the design of artificial protein compartments as nanoreactors with tunable porosity and function.

Pore structure controls stability and molecular flux in engineered protein cages.

Protein cages are a common architectural motif used by living organisms to compartmentalize and control biochemical reactions. While engineered protein cages have featured in the construction of nanoreactors and synthetic organelles, relatively little is known about the underlying molecular parameters that govern stability and flux through their pores. In this work, we systematically designed 24 variants of the Thermotoga maritima encapsulin cage, featuring pores of different sizes and charges. Twelve pore variants were successfully assembled and purified, including eight designs with exceptional thermal stability. While negatively charged mutations were better tolerated, we were able to form stable assemblies covering a full range of pore sizes and charges, as observed in seven new cryo-EM structures at 2.5- to 3.6-Å resolution. Molecular dynamics simulations and stopped-flow experiments revealed the importance of considering both pore size and charge, together with flexibility and rate-determining steps, when designing protein cages for controlling molecular flux.

The iron islands: Erythroblastic islands and iron metabolism.

BACKGROUND: A healthy human can produce over 1 × 1015 blood cells throughout their life. This remarkable amount of biomass requires a concomitantly vast amount of iron to generate functional haemoglobin and functional erythrocytes. SCOPE OF THE REVIEW: Erythroblasts form multicellular clusters with macrophages in the foetal liver, bone marrow and spleen termed erythroblastic islands. How the central erythroblastic island macrophage co-ordinates the supply of iron to the developing erythroblasts will be a central focus of this review. MAJOR CONCLUSION: Despite being studied for over 60 years, the mechanisms by which the erythroblastic island niche serves to control erythroid cell iron metabolism are poorly resolved. GENERAL SIGNIFICANCE: Over 2 billion people suffer from some form of anaemia. Iron deficiency anaemia is the most prevalent form of anaemia. Therefore, understanding the processes by which iron is trafficked to, and metabolised in developing erythrocytes, is crucially important.