Biology subjects

Lei, D.

Publications and source records attributed to Lei, D..

8 recordsLinked to original sources

bioRxiv2026

Structural basis for repurposing a flexible phage tail into an Intraspecific bacterial competition weapon

The rise of multidrug-resistant bacteria poses a major threat to human health. Tailocins, which are phage tail-like bacteriocins, offer a way to kill specific bacteria without the risks of horizontal gene transfer found in phage therapy. While the mechanism of contractile R-type tailocins is well-established, the bactericidal action of non-contractile F-type tailocins remains unresolved. Here, we report the high-resolution cryo-EM structure of the F-pyocin from Pseudomonas aeruginosa. The structure reveals a complex nanomachine made of a terminator cap (AlpD/PA0910), a helical tube (PA0633), and a tail tip that mediates a symmetry transition from sixfold to threefold. Notably, the tail tip connects to an elongated central fiber (PA0641) decorated with trimeric side fibers (PA0643/PA0646) involved in host recognition. Through genetic deletion studies and proteomic analysis, we delineate the F-pyocin assembly pathway, establishing that the tape-measure protein (PA0636) serves as a central scaffold coordinating the assembly of the tube, cap, and tail-tip modules. Structural analysis reveals a localized register shift in the central fibers coiled-coil shaft, suggesting a metastable state that may facilitate conformational changes upon host recognition. Our findings detail the atomic structure of an F-type tailocin, uncovering its coordinated assembly pathway and specialized tail-fiber architecture, which establishes a structural framework for engineering next-generation, precision antimicrobials.

biophysics↗

bioRxiv2025

Pulmonary mRNA-LNP Vaccines for Rapid and Durable Protection Against Bacterial Infection

Pulmonary bacterial infections remain a major clinical challenge. Although vaccination reduces infection rates and mortality, the vulnerable post-vaccination immunity gap can still result in infection and vaccine failure. In addition, effective vaccines are unavailable for many clinically important bacterial pathogens. Here, we report a pulmonary mRNA-lipid nanoparticle (mRNA-LNP) vaccine incorporating a novel ionizable lipid engineered for localized high-level expression, which elicits both rapid and durable protections against bacterial lung infections, effectively bridging this critical window of vulnerability. Intratracheal delivery of mRNA-LNP rapidly primes lung neutrophils and macrophages into a transcriptionally pre-activated state, enhancing their phagocytic activity and enabling rapid, antigen-independent bacterial clearance during the early post-vaccination period (approximately 1-7 days). Subsequently, vaccination induces potent antigen-specific adaptive responses, conferring sustained protection against both laboratory and clinical drug-resistant Pseudomonas aeruginosa strains. Single-cell transcriptomics and immune profiling reveal coordinated activation of innate and adaptive immune programs. This dual-phase immune response exemplifies a paradigm-shifting vaccine design that integrates innate and adaptive immunity to confer both immediate and long-term protection. Our findings establish a mechanistic basis for rapid antibacterial defense and highlight pulmonary mRNA-LNP vaccination as a promising strategy for combating respiratory infections.

bioengineering↗

bioRxiv2025

Iterative acylation on mature lasso peptides by widespread acetyltransferases for lipolasso production

The biosynthesis of ribosomally synthesized and post-translationally modified peptides (RiPPs) leverages iterative catalysis to enhance structural and biological diversity. Traditionally, iterative enzymes install multiple post-translational modifications (PTMs) on linear peptides, rather than mature RiPPs with intricate three-dimensional structures, which would require complex changes in substrate binding poses. Here, we present a prolific class of GCN5-related N-acetyltransferases (GNATs) that iteratively and consecutively acylate two Lys residues within the loop and ring motifs of lasso peptides, diverging from the typical iterative modification of linear peptides--an unprecedented function for PTM enzymes. Utilizing high-resolution cryo-electron microscopy and enzymatic reconstitution, we mapped the lasso peptide binding pocket of IatT and pinpointed key residues involved in demarcating the two distinct acetylation steps. Structure-based engineering of IatTs acetyl group recognition site expanded the cavity to accommodate longer-chain acyl groups, leading to the creation of lipolasso peptides, a novel class of ribosomal lipopeptide. This engineering strategy can be applied to any RiPP BGC encoding GNAT, facilitating the efficient diversification of rare ribosomal lipopeptides. Graphic Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=95 SRC="FIGDIR/small/630886v1_ufig1.gif" ALT="Figure 1"> View larger version (30K): org.highwire.dtl.DTLVardef@1d88af9org.highwire.dtl.DTLVardef@18c1298org.highwire.dtl.DTLVardef@118b9f3org.highwire.dtl.DTLVardef@580a50_HPS_FORMAT_FIGEXP M_FIG C_FIG

synthetic biology↗

bioRxiv2021

Elevated islet prohormone ratios as indicators of insulin dependency in islet transplant recipients

Autologous pancreatic islet transplantation is an established therapy for patients with chronic pancreatitis. However, the long-term transplant outcomes are modest. Identifying indicators of graft function will aid the preservation of transplanted islets and glycemic control. To this end, we analyzed beta cell prohormone peptide levels in a retrospective cohort of total pancreatectomy autologous islet transplant patients (n=28). Proinsulin-to-C-peptide (PI/C) and proIAPP-to-total IAPP (proIAPP/IAPP) ratios measured at 3 months post-transplant were significantly higher in patients who remained insulin dependent at 1 year follow-up. In a mouse model of human islet transplantation, recipient mice that later became hyperglycemic displayed significantly higher PI/C ratios than mice that remained normoglycemic. Histological analysis of islet grafts showed reduced insulin- and proinsulin-positive area, but elevated glucagon-positive area in grafts that experienced greater secretory demand. Increased prohormone convertase 1/3 was detected in glucagon-positive cells, and glucagon-like peptide 1 (GLP-1) area was elevated in grafts from mice that displayed hyperglycemia or elevated plasma PI/C ratios, demonstrating intra-islet incretin production in metabolically challenged human islet grafts. These data indicate that in failing grafts, alpha cell prohormone processing is likely altered, and incomplete beta cell prohormone processing may be an early indicator of insulin dependency.

pathology↗

bioRxiv2020

MEMS directed evolution of two cytochrome P450 enzymes revealing distinct active-sites for convergent function

Directed evolution (DE) inspired by natural evolution (NE) has been achieving tremendous successes in protein/enzyme engineering. However, the conventional one-protein-for-one-task DE cannot match the multi-proteins-for-multi-tasks NE in terms of screening throughput and efficiency, thus often failing to meet the fast-growing demands for biocatalysts with desired properties. In this study, we design a novel multi-enzyme-for-multi-substrate (MEMS) DE model and establish the proof-of-concept by running a NE-mimicking and higher-throughput screening on the basis of two-P450s-against-seven-substrates (2Px7S) in one pot. With the significantly improved throughput and hit-rate, we witness a series of convergent evolution events of the two archetypal cytochrome P450 enzymes (P450 BM3 and P450cam) in laboratory. Further structural analysis of the two functionally convergent P450 variants provide important insights into how distinct active-sites can reach a common catalytic goal.

biochemistry↗

bioRxiv2019

Gene pathogenicity prediction of Mendelian diseases via the Random Forest algorithm

The study of Mendelian diseases and the identification of their causative genes are of great significance in the field of genetics. The evaluation of the pathogenicity of genes and the total number of Mendelian disease genes are both important questions worth studying. However, very few studies have addressed these issues to date, so we attempt to answer them in this study. We calculated gene pathogenicity prediction (GPP) score by a machine learning approach (random forest algorithm) to evaluate the pathogenicity of genes. When we applied the GPP score to the testing gene set, we obtained accuracy of 80%, recall of 93% and area under the curve (AUC) of 0.87. Our results estimated that a total of 10,399 protein-coding genes were Mendelian disease genes. Furthermore, we found the GPP score was positively correlated with the severity of disease. Our results indicate that GPP score may provide a robust and reliable guideline to predict the pathogenicity of protein-coding genes. To our knowledge, this is the first trial to estimate the total number of Mendelian disease genes.

genetics↗

bioRxiv2018

3D images of Neuronal Adhesion Molecule Contactin-2 Reveal an Unanticipated Two-State Architecture

Contactins (CNTNs) are important cell adhesion molecules that mediate neuronal and axoglial contacts, and lesions in these molecules are linked to neuropsychiatric disorders. The extracellular domain of CNTNs contains six Ig domains and four FNIII domains. Crystal structures have shown that Ig1-Ig4 forms a horseshoe-shaped headpiece, in which the N-terminal domains might fold back on the C-terminal domains to form molecular super-U shaped architecture. The arrangement of these domains has been controversial, which may due to the structural dynamics and conformation heterogeneity of the protein. Here, we used a single-molecule 3D imaging method, individual-particle electron tomography (IPET), to study the extracellular domain of CNTN2 that forms monomers with a broad spectrum of conformations, and obtained 60 three-dimensional (3D) reconstructions. In addition to the known horseshoe-shaped headpiece, ~75% headpieces unexpectedly adopt an open (elongated) or a semi-open conformations contributed to our understanding about structural dynamics. The ectodomains formed curve but not double-back in any uniform way, with an averaged molecular dimension of ~255 [A]. The first-time demonstration of the dynamic nature and conformational preferences of the full-length CNTN2 ectodomain suggest that the headpiece exists in equilibrium in the closed or not-closed states. The important architecture may provide a structural platform for protein partners to influence this balance regulating the function of CNTN2. Encoding the ability of this neural adhesion molecule to form both homomers with itself, as well as recruit different protein partner to neuronal and axoglial contact points play the key role in mediating cell-cell interactions.

biochemistry↗

bioRxiv2018

Neuroglial Remodeling After Various Ocular Injuries is Regulated by Microglia

Reactive microglia and infiltrating peripheral monocytes have been implicated in many neurodegenerative diseases of the retina and central nervous system (CNS). However, their specific contribution in retinal degeneration remains unclear. We recently showed that peripheral monocytes that infiltrate the retina after ocular injury in mice become permanently engrafted into the tissue, establishing a pro-inflammatory phenotype that promotes neurodegeneration. Here, we show in mice that microglia regulate the process of neuroglia remodeling during ocular injury, and their depletion results in marked upregulation of inflammatory markers, such as Il17f, Tnfsf11, Ccl4, Il1a, Ccr2, Il4, Il5, and Csf2 in the retina, abnormal engraftment of peripheral CCR2+ CX3CR1+ monocytes into the retina and is associated with increased retinal ganglion cell (RGC) loss, retinal nerve fiber layer thinning, and RPE65+ cell migration onto the retinal surface. Furthermore, we show that other types of ocular injuries, such as penetrating corneal trauma and ocular hypertension, also cause similar changes. However, optic nerve crush injury mediated RGC loss evokes neither peripheral monocyte response in the retina, nor RPE65+ cell migration, although peripheral CX3CR1+ and CCR2+ monocytes infiltrate the optic nerve injury site and remain present for months. Our study suggests that microglia are key regulators of peripheral monocyte infiltration and RPE migration and their depletion results in abnormal neuroglia remodeling that exacerbates neuroretinal tissue damage. This mechanism of retinal damage through neuroglia remodeling may be clinically important for the treatment of patients with ocular injuries, including surgical traumas.

immunology↗