Biology subjects

Yan, C.

Publications and source records attributed to Yan, C..

At least 37 records · Page 2Linked to original sources

bioRxiv2025

Real-Time Viability Assessment of Ex Vivo Mouse Kidneys for Transplant Applications Using Dynamic Optical Coherence Tomography

Kidney transplantation remains the preferred treatment for patients with end-stage kidney disease. However, the ongoing shortage of donor organs continues to limit the availability of transplant treatments. Existing evaluation methods, such as the kidney donor profile index (KDPI) and pre-transplant donor biopsy (PTDB), have various limitations, including low discriminative power, invasiveness, and sampling errors, which reduce their effectiveness in organ quality assessment and contribute to the risk of unnecessary organ discard. In this study, we explored the dynamic optical coherence tomography (DOCT) as a real-time, non-invasive approach to monitor the viability of ex vivo mouse kidneys during static cold storage over 48 hours. The dynamic metrics logarithmic intensity variance (LIV), early OCT correlation decay speed (OCDSe), and late OCT correlation decay speed (OCDSl) were extracted from OCT signal fluctuations to quantify temporal and spatial tissue activity and deterioration. Our results demonstrate that DOCT provides complementary functional information to morphological assessment offered by conventional OCT imaging, showing potential to improve pre-transplant organ evaluation and clinic decision-making.

bioengineering↗

bioRxiv2025

Methionine γ-lyase Traps Cofactor Pyridoxal-5'-phosphate through Specific Serine-Mediated Affinity

Pyridoxal-5'-phosphate (PLP), the ubiquitous and ancient cofactor, plays important roles in enzymatic elimination, transamination and other reactions. The catalytic efficiency of PLP-dependent enzymes is significantly higher than that of free PLP. The recruitment of PLP from the environment by the enzymes, particularly through interactions outside the active site, is the key step determining the occurrence of PLP-mediated catalysis. However, the precise mechanism by which enzymes recruit PLP remains elusive. Methionine {gamma}-lyase (MGL), a PLP-dependent enzyme, catalyzes the degradation of L-methionine, thereby suppressing cancer cell proliferation through serum or dietary methionine depletion. Here, we report the crystal structure of yMGL, which belongs to a newly identified subgroup of cystathionine {gamma}-lyases, in complex with L-methionine and PLP. Through truncating the C-terminal domain of yMGL both in vitro and in vivo, we demonstrated that this domain, outside the canonical PLP-binding domain, is essential for the specific interaction between yMGL and PLP, as well as for efficient L-methionine catabolism. A conserved Ser in the C-terminal domain, positioned at the active-site entrance of MGLs, confers specificity for PLP. These findings elucidate a previously uncharacterized mechanism of PLP recruitment by MGLs and promote rational design of MGLs with tailored cofactor selectivity and catalytic performance.

biochemistry↗

bioRxiv2025

Mechanism-Aware Protein-Protein Interaction Prediction via Contact-Guided Dual Attention on Protein Language Models

Robust prediction of protein-protein interactions (PPIs) requires models that generalize beyond the training distribution. Here, we present PLMDA-PPI, a mechanism-aware framework that incorporates biophysical inductive biases into a dual-attention architecture with residue-level contact supervision applied to protein language model embedded geometric representations. This design enables the model to jointly learn global interaction likelihoods and the residue pairs mediating interactions, producing interpretable, mechanism-grounded predictions. PLMDA-PPI demonstrates strong out-of-distribution generalization, substantially outperforming lightweight deep learning models (D-SCRIPT, Topsy-Turvy, TT3D, and TUnA) and achieving comparable or even superior performance to computationally intensive methods (AF2Complex, RF2-Lite, and RF2-PPI) derived from AlphaFold2 and RoseTTAFold2, while requiring orders of magnitude fewer computational resources. These results show that incorporating mechanistic priors as architectural inductive biases enhances generalization, interpretability, and computational efficiency, providing a principled foundation for AI-driven prediction of complex biological interactions.

bioinformatics↗

bioRxiv2025

Adaptation to warm environments with a fast pace of life in a marine predatory snail

Understanding how latitudinal temperature variation shapes local adaptation of life history strategies is crucial for predicting future responses to warming. Contrasting predictive frameworks explain how growth and other life history traits may respond to differing selective pressures across latitude. However, these frameworks have rarely been explored within the context of fluctuating environmental temperatures across longer (i.e., seasonal) time scales experienced in nature. Furthermore, consequences of growth differences for other aspects of fitness, including reproductive output, remain unclear. Here, we conducted a long-term (17-month) simulated reciprocal transplant experiment to examine local adaptation in two populations of the predatory marine snail Urosalpinx cinerea separated by 8.6{degrees} latitude (1000 km). We reared F1 offspring under two seasonally fluctuating temperature regimes ("warm" and "cold", simulating field thermal conditions experienced by low and high latitude populations, respectively), quantifying temporal patterns in growth, maturation, and reproductive output. We identified striking divergence in life-history strategies between populations in the warm regime, with offspring from the low latitude population achieving greater growth in their first year, and high reproductive output coupled with reduced growth in their second year. In contrast, the high latitude population grew slower in their first year, but eventually attained larger sizes in their second year, at the expense of reduced reproductive output. Responses were consistent with this in the cold regime, although growth and reproductive output was reduced in both populations. Our data provides support for adaptive divergence across latitude consistent with the Pace-of-Life hypothesis, with the low latitude population selected for a fast-paced life characterized by rapid development and early reproduction. In contrast, the high latitude population exhibited slower growth and delayed maturation. Our results highlight the potential limitations of short-term comparisons of growth without considering processes over longer time scales that may exhibit seasonal temperature variation and ontogenetic shifts in energy allocation and imply a radical reshaping of physiological performance and life history traits across populations under climate change.

ecology↗

bioRxiv2025

Identifying Key Hub Genes that Attribute Varying Host Responses: A Longitudinal RNA-seq Analysis of SARS-CoV-2 Delta and Omicron Infections.

Coronavirus disease 2019 (COVID-19), caused by SARS-CoV-2, has severely impacted global health, with different Variants of Concern (VOCs) resulting in varied clinical outcomes. Among these, the Delta and Omicron variants have drawn significant attention due to their distinct characteristics--Delta is associated with higher virulence and Omicron has greater transmissibility but reduced severity. This study compares host immune responses to the Delta and Omicron variants using RNA-seq data from infected primary human airway epithelial cells. Both variants triggered a robust antiviral innate immune response by 2 days post-infection (dpi). However, Omicron was found to elicit a more rapid immune response, showing pathway enrichment at 1 dpi. In contrast, Delta displayed no immune-related pathway activation within the first 24 hours, suggesting it may evade early immune detection, promoting increased viral replication. By 3 dpi, Delta induced a more aggressive immune response, particularly in pathways related to cell death and pro-inflammatory signaling, such as "programmed cell death" and "regulation of cell death." Weighted gene co-expression network analysis revealed distinct immune-related genes: Delta infections were characterized by hub genes like MYD88 and IL1R1 involved in pro-inflammatory responses, HLA-A & B, NLRC5 and PSMB9 involved in antigen presentation and TNSF10 and IFR1 involved in pro-apoptotic processes. Conversely, Omicron infections were marked by hub genes such as CXCL1 and CXCL8 involved in immune cells recruitment, MET and LYN involved in reducing hyperinflammatory responses and maintaining immune balance, and IFI44, EIF2AK2 and IFIT5 responsible for the sensing of viral RNA among others.

bioinformatics↗

bioRxiv2025

Student-led experimental evolution reveals novel biofilm regulatory networks underlying adaptations to multiple niches

We established a research-education partnership, EvolvingSTEM, that currently provides thousands of secondary school students the opportunity to conduct authentic research experiments centered on microbial evolution each year. Providing high school students access to research experiences not only improves learning and can have positive and long-lasting impacts on their attitudes towards science, but also gives them the opportunity to make impactful scientific contributions. Through EvolvingSTEM, students evolve populations of Pseudomonas fluorescens in a bead model that includes daily cycles of bacterial dispersal, attachment, and biofilm growth and observe heritable changes in colony morphology. Genome sequencing of 70 mutants that they picked identified parallel mutations in genes known to regulate biofilm growth (wsp, yfiBNR, morA, fuzY) and uncovered novel adaptations: loss-of-function mutations in phosphodiesterase PFLU0185 that did not alter colony morphology and mutations affecting periplasmic disulfide bond formation producing small colonies. PFLU0185 mutants rapidly and consistently reached high frequencies and phenotyping revealed roles in cyclic di-GMP regulation, biofilm formation, and motility, prompting us to name this gene bmo (biofilm and motility regulator). Competition experiments and microscopy demonstrated bmo mutants employ generalist strategies and coexist with their ancestor and specialist mutants through niche differentiation. Consequently, phenotypic diversity is maintained, with smooth (ancestral and bmo) colonies consistently outnumbering wrinkly and fuzzy variants. This study advances our understanding of biofilm genetic architecture while demonstrating that student-led research can uncover mechanisms of microbial adaptation relevant to Pseudomonas infection biology. IMPORTANCEBacterial biofilms dominate microbial life, yet their evolutionary genetics remain incompletely understood. Extensive replication of experiments that employ similar, but not identical, biofilm selection models can provide valuable insights into mechanisms of adaptation. We demonstrate that this can be achieved through university-education partnerships that engage secondary school students in authentic research. Student-led experiments revealed that loss-of-function mutations in a conserved phosphodiesterase, PFLU0185/bmo, dominate evolved populations without changing colony morphology. This finding, combined with diverse, less frequent mutants that alter colony morphology informs the process of biofilm niche differentiation. This work also demonstrates the power of distributed research networks for discovering new genetic pathways of adaptation. Students gained authentic research experience, potentially inspiring them to become scientists, while identifying mutants adapted to discrete conditions that maintain diversity within biofilms. This synergy between education and discovery offers a scalable model for addressing complex biological questions while developing scientific literacy in diverse classrooms.

microbiology↗

bioRxiv2025

Stick-slip motion and universal statistics of cargo transport within living cells

Cargo transport within cells is a vital biological process that relies on the intricate interplay between motor proteins, microtubules, and the complex intracellular environment. In this study, we unveil a universal transport mechanism characterized by stick-slip motion, which governs the dynamics of intracellular vesicle transport. By analyzing a comprehensive dataset of vesicle trajectories across various cell types and intracellular environments, we demonstrate that the cargo velocities consistently follow a Gamma distribution, revealing a common statistical pattern amidst the diversity of biological cargoes. Our experimental findings are well-described by a theoretical model that connects the Brownian-correlated kinetic friction between motor-cargo complexes and their surroundings to the observed universal Gamma distribution of cargo velocities. This model elucidates the stick-slip dynamics governing intracellular cargo transport, which are pertinent to various cellular processes such as vesicle budding, organelle transport, and cell migration.

biophysics↗

bioRxiv2025

The interaction of XPG with TFIIH through p62 and XPD is required for the completion of nucleotide excision repair

Nucleotide excision repair (NER) is the key pathway for the removal of DNA damage induced by UV irradiation and chemotherapeutic reagents. Protein-protein interactions are crucial for the dynamic and coordinated assembly of the proteins involved at DNA lesion. Here we focus on the role of interactions between the multi-subunit helicase/translocase complex TFIIH and the 3 endonuclease XPG. We show that XPG interacts with the p62 and XPD subunits of TFIIH through its long spacer region located in the middle of its split active site. We show the interactions between three acidic regions of XPG with the pleckstrin homology (PH) domain of p62 are of moderate importance for NER, while interactions with XPD are critical for activity. Combining mutations in the p62 and XPD interaction domains leads to additive defects in NER activity. Unexpectedly, we show that these interactions are not required for the recruitment of XPG, but rather for the formation of a catalytically competent NER complex and for triggering the incision 5 to the lesion by ERCC1-XPF. Our studies provide fundamental insights into how interactions between TFIIH and XPG contribute to the NER pathway and more generally how modular protein-protein interactions control each step along the NER reaction coordinate.

biochemistry↗

bioRxiv2025

Upstream open reading frame inactivation augments GATA4 translation and cardiomyocyte hypertrophy in mice

Upstream open reading frames (uORFs) are short peptide-encoding sequences located in the 5 untranslated region (5 UTR) of mRNAs, enabling translational repression of main (m)ORFs. While uORFs are found in [~]50% of mRNAs in humans, our understanding of their biological function remains limited. This study aims to elucidate the role of the uORF in the 5 UTR of the Gata4 (GATA binding protein 4) gene in cardiac biology by inactivating its start codon ({Delta}uORF) in the mouse genome. Our investigation reveals that mice with Gata4 uORF inactivation manifest spontaneous cardiac hypertrophy without apparent fibrosis as they age. Utilizing single-nucleus RNA sequencing (snRNA-seq), we uncovered significant transcriptional variations between wild-type (WT) and {Delta}ORF mice. Notably, mRNAs associated with sarcomeres and contractile functions show heightened expression levels, reflecting the hypertrophic phenotype. Notably, at least nine upregulated genes are GATA4-bound targets in mouse ventricles. Functional assessments of isolated primary adult cardiomyocytes confirmed enhanced hypertrophy and contractility in {Delta}ORF mice. Additionally, we employed single-nucleus transposase-accessible chromatin (snATAC)-seq to investigate changes in chromatin accessibility. Our results indicated increased accessibility within specific transcription-regulatory elements linked to elevated gene transcription. These putative cis-regulatory elements (pCREs) are significantly enriched in MEF2 (myocyte enhancer factor 2) binding motifs. In vitro luciferase reporter assays further supported the regulatory potential of three of these pCREs, highlighting their role in the transcriptional enhancement of three GATA4 target genes bound by MEF2 and GATA4. These findings illuminate the role of uORF in negatively regulating GATA4 protein expression and cardiomyocyte hypertrophy at the organismal level and provide a novel therapeutic target for cardiac pathogenesis.

biochemistry↗

bioRxiv2025

AI-driven computational pathology for recurrence risk assessment in early-stage breast cancer

Recurrence related to poor prognosis is a leading cause of mortality in patients with breast cancer (BC). The MammaPrint (MP) genomic assay is designed to stratify recurrence risk and evaluate chemotherapy benefits for early-stage HR+/HER2- BC patients. However, MP fails to reveal spatial tumor morphology and is limited by high costs. In this study, we establish a BC MP cohort and develop CPMP, a weakly supervised agent-attention transformer model, to predict MP recurrence risk from annotation-free BC histopathological slides. CPMP achieves an AUROC of 0.824 {+/-} 0.03 in predicting MP risk groups. We further leveraged CPMP for spatial and morphological analyses to explore histological patterns associated with MP risk groups. The model reveals tumor spatial localization at the whole-slide level and highlights distinct intercellular interaction patterns of MP groups. It also characterizes the diversity in tumor morphology and uncovers MP High-specific, Low-specific, and colocalized morphological phenotypes that differ in quantitative cellular composition. Prognostic evaluation in the external cohort exhibits significant stratification of distant metastasis risk (HR: 3.14, p-value = 0.0014), underscoring the prognostic power of CPMP. These findings demonstrate the capability of CPMP in MP risk prediction, offering a flexible supplement to genomic risk assessment in early-stage BC.

pathology↗

bioRxiv2025

A pan-positive allosteric modulator that mediates sustainable GPCR activation

Approximately 1/3 of all clinical drugs exert their therapeutic effects by modulating the activity of G protein-coupled receptors (GPCRs). Thus, there is a constant interest in finding novel ways to modulate GPCR activity. In this work, through a newly established Survival Pressure Selection (SPS) method for high-throughput screening of GPCR agonists, we discover that atazanavir functions as an agonist for GPR119. Further studies suggest that atazanavir is capable of activating a number of Family A GPCRs, including the {beta}1 adrenergic receptor ({beta}1AR), the {beta}2 adrenergic receptor ({beta}2AR) and the opoioid receptor (OR). Cryo-EM structures reveal that atazanavir binds to an allosteric pocket near TM6/7 in both GPR119 and the {beta}1AR. Pharmacological studies suggest that atazanavir mediates non-canonical signaling of Family A GPCRs. During this process, G protein and {beta}-arrestin are spatially close, but {beta}-arrestin forms a complex with the receptor and G protein instead of desensitizing G protein signaling, thus resulting in sustained G protein signaling. The work expands the signal transduction modes and pharmacological properties of allosteric modulators for GPCRs and provides a general starting point for developing therapeutics targeting this allosteric site.

biochemistry↗

bioRxiv2025

A GPCR-G protein-β-arrestin megacomplex induced by an allosteric modulator

GPCRs signal through both G protein pathways and {beta}-arrestin pathways. Previous work suggests that {beta}-arrestins bind to GPCRs through different modes, including core engagement and tail engagement. Core engagement competes with G proteins and terminates G protein signaling, while tail engagement can coexist with G proteins, mediating sustained intracellular activation of the receptor - a process dependent on the high affinity between {beta}-arrestin and the phosphorylated C-terminus of the receptor. In this study, we determined the structure of a GPCR - G protein - {beta}-arrestin-1 complex stabilized by an allosteric modulator. The compound, atazanavir, acts like molecular glue to anchor {beta}-arrestin-1 to the receptors TM6 and TM7 regions. This pendulum binding mode is structurally compatible with simultaneous G protein binding. We further demonstrate that the atazanavir-mediated {beta}-arrestin recruitment does not require the receptors C-terminal region. This work illustrates a novel paradigm of GPCR-G protein-{beta}-arrestin1 megacomplex assembly and opens up new avenues for modulating GPCR function.

biochemistry↗

bioRxiv2025

A rapidly prototyped, simple yet versatile dynamic breathing Exposure-on-a-Chip for investigating nanoparticle deposition in the alveoli

We developed and characterized a three-layer microfluidic Exposure-on-a-Chip (EOC) model to replicate the alveolar microenvironment and simulate breathing motions, offering a physiologically relevant platform for studying inhaled nanomedicines. The EOC chip, fabricated from biocompatible polydimethylsiloxane (PDMS), features a fluidic chamber for cell culture, a pneumatic chamber for pressure application, and a thin PDMS membrane (150 {micro}m) separating the chambers to support cell growth and enable mechanical stretching. Xurography and 3D printing were validated as efficient and reproducible fabrication methods. Mechanical characterization, using fluorescent bead tracing, confirmed that the PDMS membrane accurately mimics alveolar breathing motions under physiological conditions (1-8% strain). Biological validation showed that alveolar epithelial and endothelial cells cultured on the EOC formed functional monolayers, maintaining barrier integrity under cyclic and static stretching, separately. To study nanoparticle behavior, we examined the deposition of nanoparticles under dynamic stretching versus static conditions. Significantly fewer nanoparticles accumulated in cells under continuous dynamic exposure with stretching compared to static culture, highlighting the critical role of mechanical forces in nanoparticle-cell interactions. The EOC platform provides a robust and scalable tool for evaluating nanomedicine efficacy in dynamic environments, representing a significant advancement in alveolus-on-chip technology.

bioengineering↗

bioRxiv2025

Themis dominates T cell exhaustion by regulation of TCR and PD-1 signaling

T cell exhaustion is important to protect the host from immunopathology during chronic viral infection, but it also impairs T cell anti-tumor immunity1-5. A fundamental unresolved question is whether and how T cell exhaustion is determined at the onset of TCR signaling6-8. Here we report an unexpected role of Themis, a TCR-proximal signaling molecule9, in T cell exhaustion. Chronic viral infection in mice usually leads to T cell exhaustion and survival of the host. Surprisingly, Themis T-cell conditional knockout mice died from severe CD8+-dependent lung immunopathology in chronic viral infection, showing Themis importance in establishing T cell exhaustion. We found that Themis-deficient CD8+ T cells were hyperactivated at the single-cell level - producing more TNF and IFN{gamma} compared to wild-type counterparts - but defective in population-level expansion. Moreover, TCF-1 and TOX expression were inhibited in Themis-deficient CD8+ T cells, thereby impairing differentiation of exhausted T cell precursors (T-pex) and maintenance of terminally exhausted T cells (T-ex), respectively. Mechanistically, Themis initially promotes TCR signaling to induce PD-1 expression and subsequently mediates PD-1 signaling. In the latter, Themis binds to PD-1 and promotes PD-1 phosphorylation and its recruitment of SHP2, thereby acting as a negative regulator to inhibit T cell effector functions. Without Themis, the orderly regulation of TCR and PD-1 signaling, and therefore exhaustion, is disrupted. Thus, our results unequivocally demonstrate that Themis-mediated early TCR signaling plays a decisive role in T cell exhaustion and provide a novel mechanism of PD-1 signaling through Themis.

immunology↗

bioRxiv2025

Amyloid-β precursor protein promotes tumor growth by establishing an immune-exclusive tumor microenvironment

During initiation and progression, cancerous tissue hijacks a series of elaborate tissue homeostatic mechanisms to avoid immune surveillance, including neuro-immune interactions. Here, we show that amyloid-{beta} precursor protein (APP) and its {beta}-cleavage product amyloid-{beta}1-42 (A{beta}1-42), well-known in the pathogenesis of Alzheimers disease (AD), are expressed in multiple cancer tissues. However, the oncogenic activity of APP is due to its E1 domain, instead of A{beta}1-42. Mechanistically, APP restricts immune cells influx into tumor microenvironment (TME) and impairs CD8+ T cell and NK cell-based immunity, by dampening type I interferon (IFN) response in TME. We also provide proof-of-concept that vaccination targeting APP is effective for cancer prevention. Our current study reveals a previously unrecognized role of APP in cancer immune surveillance, and provides a new strategy for cancer prevention and treatment by targeting APP.

cancer biology↗

bioRxiv2025

Neoadjuvant anti-4-1BB confers protection against spontaneous metastasis through low-affinity intratumor CD8+ T cells in triple-negative breast cancer

Neoadjuvant immunotherapy seeks to harness the primary tumor as a source of relevant tumor antigens to enhance systemic anti-tumor immunity through improved immunological surveillance. Despite having revolutionized the treatment of patients with high-risk early-stage triple-negative breast cancer (TNBC), a significant portion of patients remain unresponsive and succumb to metastatic recurrence post-treatment. Here, we found that optimally scheduled neoadjuvant administration of anti-4-1BB monotherapy was able to counteract metastases and prolong survival following surgical resection. Phenotypic and transcriptional profiling revealed enhanced 4-1BB expression on tumor-infiltrating intermediate (Tint), relative to progenitor (Tprog) and terminally exhausted (Tterm) T cells. Furthermore, Tint was enriched in low-affinity T cells. Treatment with anti-4-1BB drove clonal expansion of Tint, with reduced expression of tissue-retention marker CD103 in Tprog. This was accompanied by increased TCR clonotype sharing between paired tumors and pre-metastatic lungs. Further interrogation of sorted intratumor T cells confirmed enhanced T cell egress into circulation following anti-4-1BB treatment. In addition, gene signature extracted from anti-4-1BB treated Tint was consistently associated with improved clinical outcomes in BRCA patients. Combinatorial neoadjuvant anti-4-1BB and ablation of tumor-derived CXCL16 resulted in enhanced therapeutic effect. These findings illustrate the intratumor changes underpinning the efficacy of neoadjuvant anti-4-1BB, highlighting the reciprocity between local tissue-retention and distant immunologic fortification, suggesting treatment can reverse the siphoning of intratumor T cells to primary tumor, enabling redistribution to distant tissues and subsequent protection against metastases.

immunology↗

bioRxiv2025

Triple-effect correction for Cell Painting data with contrastive and domain-adversarial learning

Cell Painting (CP), as a high-throughput imaging technology, generates extensive cell-stained imaging data, providing unique morphological insights for biological research. However, CP data contains three types of technical effects, referred to as triple effects, including batch effects, gradient-influenced row and column effects (well position effects). The interaction of various technical effects can obscure true biological signals and complicate the characterization of CP data, making correction essential for reliable analysis. Here, we propose cpDistiller, a triple-effect correction method specially designed for CP data, which leverages a pre-trained segmentation model coupled with a semi-supervised Gaussian mixture variational autoencoder utilizing contrastive and domain-adversarial learning. Through extensive qualitative and quantitative experiments across various CP profiles, we demonstrate that cpDistiller effectively corrects triple effects, especially well position effects, a challenge that no current methods address, while preserving cellular heterogeneity. Moreover, cpDistiller effectively captures system-level phenotypic responses to genetic perturbations and reliably infers gene functions and interactions both when combined with scRNA-seq data and independently. cpDistiller also excels at identifying gene and compound targets, which is a critical step in drug discovery and broader biological research.

bioinformatics↗

bioRxiv2024

High-resolution mastigoneme structure reveals 5',5'-phosphodiesters stabilized glycan folding

Glycans play a crucial role in structure organization, energy metabolism, and signal transduction in living organisms. Compared with proteins and nucleic acids, glycans exhibit remarkable molecular heterogeneity, extensive conformational flexibility, and diverse linkages. These properties pose significant challenges for obtaining high-resolution structure of glycans. Here, we report a cryo-EM structure of the heavily glycosylated Chlamydomonas mastigoneme at 2.3-2.5 [A] resolutions. In addition to enabling analysis of accurate interactions for glycosyl packing, the high resolution map reveals an unprecedented 5,5-phosphodiester bond that links adjacent glycan chains attached to hydroxyproline (Hyp) residues n and n+3. Structural analysis reveals a secondary structural element for glycoconjugates, which we name the poly-Hyp (pHP) glycohelix. Our work represents an important advancement in deciphering glycan folding.

biochemistry↗