Other Biochemistry, Biophysics, and Structural Biology Commons^™

Analysis Of The Atypical Temperature Dependence And Conformational Changes During Turnover Of A Lactobacillus Chlorogenic Acid Esterase, Nathaniel Carl, Yianni Tsigaris, Dan Ji, Nora K. Anpree, Kellie K. Omori, Cedric P. Owens

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Many bacterial chlorogenic acid esterases (ChlEs) exhibit atypical temperature behavior, featuring activities that barely change with temperature and activity maxima that fall below the thermal denaturation point. This work focuses on a ChlE from Lactobacillus helveticus (Lh–ChlE), which has a flat temperature dependence. First, it was determined that conformational changes during Lh–ChlE turnover are not rate-limiting and that the overall rate depends on the chemical step at all temperatures. Next, Lh–ChlE’s temperature dependence was investigated using a conformational equilibrium model that assumes the existence of a temperature-dependent equilibrium between an active and an inactive conformation and an activation heat capacity …

Albumin-Binding Domains In Therapeutic Protein Engineering: A Structural And Computational Perspective On Rational Design, Matthew Argyle, Dallin M. Chipman, Anna Claire Woolley, Bradley C. Bundy, Dennis Della Corte

Faculty Publications

Therapeutic proteins face a critical pharmacokinetic challenge: rapid clearance from circulation limits their clinical efficacy. Albumin-binding domains (ABDs) offer an elegant solution by enabling therapeutic proteins to “hitchhike” on serum albumin’s favorable 19-day half-life through FcRn-mediated recycling. Clinical validation through approved therapeutics like ozoralizumab demonstrates the success of this approach, with preclinical studies showing fusion to an ABD extended half-life to 18 days. This review provides an analysis of ABD-fusion protein design, integrating structural biology, computational prediction, and rational engineering principles. We catalog the major classes of albumin-binding modalities, including bacterial three-helix bundle domains, engineered peptides, antibody-derived binders, and alternative …

Benzothiazole Sulfones As A Tool For Peptide Modification And Cleavage, Phoebe R. Rubiio, Gavin A. Clausen, Zachary D. Ruighaver, Abigail F. Dalton, John P. Blobe, Amanda L. Moore, Christopher R. Shugrue

Chemistry Faculty Publications

We report the development of a new benzothiazole sulfone (BTS) tool for peptide modification and cleavage. Benzothiazole sulfones are electrophiles that rapidly react with thiols in nucleophilic aromatic substitution (SNAr) reactions. Alkyne-containing BTS compounds were incorporated onto azidolysine-containing peptides through copper-catalyzed azide-alkyne cycloadditions (CuAACs). BTS-containing peptides were found to react rapidly with two equivalents of thiol nucleophiles at room temperature in 30 min or less. BTS-containing peptides were conjugated to a variety of different compounds, including small molecules, fluorophores, and peptides. Alternatively, a BTS-based cleavable linker could also be synthesized with small molecule cargo, such as biotin, and …

Effect Of Glycation On The Binding Interactions Of Serum Albumin And Silver Nanoparticles, Megan Kirk

Celebrating Scholarship and Creativity Day (2018-)

Nanoparticles are particles ranging from 1-100 nanometers in size. Most nanoparticles, upon contact with biological matrices, are immediately coated with proteins, forming the protein corona, which can alter the structure and function of proteins. Despite the tremendous advancement of nanotechnology over the last decade, the interactions of nanoparticles with proteins are not well characterized. This project investigates the effect of glycation on the binding interactions of HSA, the most abundant protein in human blood, with silver nanoparticles using various spectroscopic techniques. This is important because in diabetic patients, HSA is exposed to high concentrations of glucose over extended periods of …

Fast 3d Printing Of Fine, Continuous, And Soft Fibers Via Embedded Solvent Exchange, Wonsik Eom, Mohammad Tanver Hossain, Vidush Parasramka, Jeongmin Kim, Ryan W. Y. Siu, Kate A. Sanders, Dakota Piorkowski, Andrew Lowe, Hyun Gi Koh, Michael F. L. De Volder, Douglas S. Fudge, Randy H. Ewoldt, Sameh H. Tawfick

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Nature uses fibrous structures for sensing and structural functions as observed in hairs, whiskers, stereocilia, spider silks, and hagfish slime thread skeins. Here, we demonstrate multi-nozzle printing of 3D hair arrays having freeform trajectories at a very high rate, with fiber diameters as fine as 1.5 µm, continuous lengths reaching tens of centimeters, and a wide range of materials with elastic moduli from 5 MPa to 3500 MPa. This is achieved via 3D printing by rapid solvent exchange in high yield stress micro granular gel, leading to radial solidification of the extruded polymer filament at a rate of 2.33 μm/s. …

Chilling- And Dark-Regulated Photoprotection In Miscanthus, An Economically Important C4 Grass, Jared Haupt, Katarzyna Glowacka

Department of Biochemistry: Faculty Publications

Tolerance of chilling dictates the geographical distribution, establishment, and productivity of C4 crops. Chilling reduces enzyme rate, limiting the sink for the absorbed light energy leading to the need for quick energy dissipation via non-photochemical quenching (NPQ). Here, we characterize NPQ upon chilling in three Miscanthus accessions representing diverse chilling tolerance in C4 grasses. High chilling tolerant accessions accumulate substantial amounts of zeaxanthin during chilling nights in both field and growth chamber settings. Chilling-induced zeaxanthin accumulation in the dark enhances rate of NPQ induction by 66% in the following morning. Based on our data, the emerging ways for the unique …

Analysis Of Natural Structures And Chemical Mapping Data Reveals Local Stability Compensation In Rna, Robert L. Cornwell-Arquitt, Riley Nigh, Michael T. Hathaway, Joseph D. Yesselman, David A. Hendrix

Department of Biochemistry: Faculty Publications

RNA molecules adopt complex structures that perform essential biological functions across all forms of life, making them promising candidates for therapeutic applications. However, our ability to design new RNA structures remains limited by an incomplete understanding of their folding principles. While global metrics such as the minimum free energy are widely used, they are at odds with naturally occurring structures and incompatible with established design rules. Here, we introduce local stability compensation (LSC), a principle that RNA folding is governed by the local balance between destabilizing loops and their stabilizing adjacent stems, challenging the focus on global energetic optimization. Analysis …

Student Comprehension Of Biochemistry In A Flipped Classroom Format, Ed Harris, Evan A. Schroder, Teryn J. Berks

Department of Biochemistry: Faculty Publications

Concept-heavy courses such as Biochemistry in life and physical science curricula are challenging for many college-aged students. It is easy for students to disengage in a lecture and not learn the subject matter while in class. To improve student learning and participation, we employed a flipped format for the first half of the course and compared learning outcomes and attitudes with the traditional lecture in the second half of the course. The experimental course was an upper-level biochemistry course taken primarily by juniors (49%), seniors (40%), and some graduate students (10%) at the college level. Our results indicate that lower-performing …

Predicting Mutation-Induced Allosteric Changes In Structures And Conformational Ensembles Of The Abl Kinase Using Alphafold2 Adaptations With Alanine Sequence Scanning, Nishank Raisinghani, Mohammed Alshahrani, Grace Gupta, Gennady M. Verkhivker

Mathematics, Physics, and Computer Science Faculty Articles and Research

Despite the success of AlphaFold2 approaches in predicting single protein structures, these methods showed intrinsic limitations in predicting multiple functional conformations of allosteric proteins and have been challenged to accurately capture the effects of single point mutations that induced significant structural changes. We examined several implementations of AlphaFold2 methods to predict conformational ensembles for state-switching mutants of the ABL kinase. The results revealed that a combination of randomized alanine sequence masking with shallow multiple sequence alignment subsampling can significantly expand the conformational diversity of the predicted structural ensembles and capture shifts in populations of the active and inactive ABL states. …

Exploring Chilling Stress And Recovery Dynamics In C4 Perennial Grass Of Miscanthus Sinensis, Karolina Sobańska, Monika Mokrzycka, Martyna Przewoźnik, Tomasz Pniewski, Katarzyna Glowacka

Department of Biochemistry: Faculty Publications

The increasing cultivation of perennial C4 grass known as Miscanthus spp. for biomass production holds promise as a sustainable source of renewable energy. Unlike the sterile triploid hybrid of M. × giganteus, which cannot reproduce through seeds, M. sinensis possesses attributes that could potentially address these limitations by effectively establishing itself through seed propagation. This study aimed to investigate how 18 genotypes of M. sinensis respond to chilling stress and subsequent recovery. Various traits were measured, including growth and biomass yield, the rate of leaf elongation, and a variety of chlorophyll fluorescence parameters, as well as chlorophyll …

Structural Analysis Of Dj-1 Glyoxalase Activity By Mix-And-Inject Serial Synchrotron Crystallography, Coleman Dolamore

Department of Biochemistry: Dissertations, Theses, and Student Research

Observing enzyme structures while they are catalyzing a reaction has been a major goal of both enzymology and structural biology for decades. With the advent of time-resolved serial X-ray crystallography using X-ray free electron lasers (XFELs) and synchrotron sources, enzyme reactions can be monitored in real time in crystallo. Here, we use two different approaches to study related enzymes involved in methylglyoxal detoxification. For human DJ-1, we used pink beam mix-and-inject serial crystallography (MISC) at the Advanced Photon Source (BioCARS 14-ID) to probe the controversial mechanism of DJ-1’s action on methylglyoxal by using serial Laue diffraction. The high flux …

Structural And Biophysical Insights Into Targeting Of Claudin-4 By A Synthetic Antibody Fragment, Satchal K. Erramilli, Pawel K. Dominik, Chinemerem P. Ogbu, Anthony A. Kossiakoff, Alex J. J.

Department of Biochemistry: Faculty Publications

Claudins are a 27-member family of ~25 kDa membrane proteins that integrate into tight junctions to form molecular barriers at the paracellular spaces between endothelial and epithelial cells. As the backbone of tight junction structure and function, claudins are attractive targets for modulating tissue permeability to deliver drugs or treat disease. However, structures of claudins are limited due to their small sizes and physicochemical properties—these traits also make therapy development a challenge. Here we report the development of a synthetic antibody fragment (sFab) that binds human claudin-4 and the determination of a high-resolution structure of it bound to claudin-4/enterotoxin complexes …

Perspectives On Computational Modeling Of Biological Systems And The Significance Of The Sysmod Community, Bhanwar Lal Puniya, Meghna Verma, Chiara Damiani, Shaimaa Bakr, Andreas Dräger

Department of Biochemistry: Faculty Publications

Motivation: In recent years, applying computational modeling to systems biology has caused a substantial surge in both discovery and practical applications and a significant shift in our understanding of the complexity inherent in biological systems.

Results: In this perspective article, we briefly overview computational modeling in biology, highlighting recent advancements such as multiscale modeling due to the omics revolution, single-cell technology, and integration of artificial intelligence and machine learning approaches. We also discuss the primary challenges faced: integration, standardization, model complexity, scalability, and interdisciplinary collaboration. Lastly, we highlight the contribution made by the Computational Modeling of Biological Systems …

Descspim: An Affordable And Easy-To-Build Light-Sheet Microscope Optimized For Tissue Clearing Techniques, Kohei Otomo, Takaki Omura, Yuki Nozawa, Steven J. Edwards, Yukihiko Sato, Yuri Saito, Shigehiro Yagishita, Hitoshi Uchida, Yuki Watakabe, Kiyotada Naitou, Rin Yanai, Naruhiko Sahara, Satoshi Takagi, Ryohei Katayama, Yusuke Iwata, Toshiro Shiokawa, Yoku Hayakawa, Kensuke Otsuka, Haruko Watanabe-Takano, Yuka Haneda, Shigetomo Fukuhara, Miku Fujiwara, Takenobu Nii, Chikara Meno, Naoki Takeshita, Kenta Yashiro, Juan Marcelo Rosales Rocabado, Masaru Kaku, Tatsuya Yamada, Yumiko Oishi, Hiroyuki Koike, Yinglan Cheng, Keisuke Sekine, Jun-Ichiro Koga, Kaori Sugiyama, Kenichi Kimura, Fuyuki Karube, Hyeree Kim, Ichiro Manabe, Tomomi Nemoto, Kazuki Tainaka, Akinobu Hamada, Hjalmar Brismar, Etsuo A. Susaki

Department of Biochemistry: Faculty Publications

Despite widespread adoption of tissue clearing techniques in recent years, poor access to suitable light-sheet fluorescence microscopes remains a major obstacle for biomedical end-users. Here, we present descSPIM (desktopequipped SPIM for cleared specimens), a low-cost ($20,000–50,000), lowexpertise (one-day installation by a non-expert), yet practical do-it-yourself light-sheet microscope as a solution for this bottleneck. Even the most fundamental configuration of descSPIM enables multi-color imaging of whole mouse brains and a cancer cell line-derived xenograft tumor mass for the visualization of neurocircuitry, assessment of drug distribution, and pathological examination by false-colored hematoxylin and eosin staining in a threedimensional manner. Academically open-sourced (https://github.com/dbsbjuntendo/ …

Multifaceted Roles Of Meg3 In Cellular Senescence And Atherosclerosis, Xiao Cheng, Mohamed Sham Shihabudeen Haider Ali, Vijaya Bhaskar Baki, Matthew Moran, Huabo Su, Xinghui Sun

Department of Biochemistry: Faculty Publications

Background and aims: Long noncoding RNAs are involved in the pathogenesis of atherosclerosis. As long non-coding RNAs maternally expressed gene 3 (Meg3) prevents cellular senescence of hepatic vascular endothelium and obesity-induced insulin resistance, we decided to examine its role in cellular senescence and atherosclerosis.

Methods and Results: By analyzing our data and human and mouse data from the Gene Expression Omnibus database, we found that Meg3 expression was reduced in humans and mice with cardiovascular disease, indicating its potential role in atherosclerosis. In Ldlr^−/− mice fed a Western diet for 12 weeks, Meg3 silencing by chemically modified …

Vitamin E Biofortification: Enhancement Of Seed Tocopherol Concentrations By Altered Chlorophyll Metabolism, Ping Qin, Peng Chen, Yuanwei Zhou, Wei Zhang, Yunyun Zhang, Jingjing Xu, Lu Gan, Yingnan Liu, Jill Romer, Peter Dörmann, Edgar B. Cahoon, Chunyu Zhang

Department of Biochemistry: Faculty Publications

Homogentisate Phytyltransferase (HPT) catalyzes condensation of homogentisate (HGA) and phytyl diphosphate (PDP) to produce tocopherols, but can also synthesize tocotrienols using geranylgeranyl diphosphate (GGDP) in plants engineered for deregulated HGA synthesis. In contrast to prior tocotrienol biofortification efforts, engineering enhanced tocopherol concentrations in green oilseeds has proven more challenging due to the integral role of chlorophyll metabolism in supplying the PDP substrate. This study show that RNAi suppression of CHLSYN coupled with HPT overexpression increases tocopherol concentrations by >two-fold in Arabidopsis seeds. We obtained additional increases in seed tocopherol concentrations by engineering increased HGA production via overexpression of …

Chromosome‑Level Assembly And Analysis Of Camelina Neglecta: A Novel Diploid Model For Camelina Biotechnology Research, Shuo Wang, Rostislav Y. Blume, Zhi‑Wei Zhou, Shaoping Lu, Tara J. Nazarenus, Yaroslav B. Blume, Weibo Xie, Edgar B. Cahoon, Ling‑Ling Chen, Liang Guo

Department of Biochemistry: Faculty Publications

Camelina neglecta is a new diploid Brassicaceae species, which has great research value because of its close relationship with the hexaploid oilseed crop Camelina sativa. Here, we report a chromosome-level assembly of C. neglecta with a total length of 210 Mb. By adopting PacBio sequencing and Hi-C technology, the C. neglecta genome was assembled into 6 chromosomes with scaffold N50 of 29.62 Mb. C. neglecta has undergone the whole-genome triplication (γ) shared among eudicots and two whole-genome duplications (α and β) shared by crucifers, but it has not undergone a specific whole-genome duplication event. By synteny analysis …

Metabolic Collaboration Between Cells In The Tumor Microenvironment Has A Negligible Effect On Tumor Growth, Johan Gustafsson, Fariba Roshanzamir, Anders Hagnestål, Sagar M. Patel, Oseeyi I. Daudu, Donald F. Becker, Jonathan L. Robinson, Jens Nielsen

Department of Biochemistry: Faculty Publications

The tumor microenvironment is composed of a complex mixture of different cell types interacting under conditions of nutrient deprivation, but the metabolism therein is not fully understood due to difficulties in measuring metabolic fluxes and exchange of metabolites between different cell types in vivo. Genome-scale metabolic modeling enables estimation of such exchange fluxes as well as an opportunity to gain insight into the metabolic behavior of individual cell types. Here, we estimated the availability of nutrients and oxygen within the tumor microenvironment using concentration measurements from blood together with a metabolite diffusion model. In addition, we developed an approach …

Emerging Roles Of Cullin-Ring Ubiquitin Ligases In Cardiac Development, Josue Zambrano-Carrasco, Jianqiu Zou, Wenjuan Wang, Xinghui Sun, Jie Li, Huabo Su

Department of Biochemistry: Faculty Publications

Heart development is a spatiotemporally regulated process that extends from the embryonic phase to postnatal stages. Disruption of this highly orchestrated process can lead to congenital heart disease or predispose the heart to cardiomyopathy or heart failure. Consequently, gaining an in-depth understanding of the molecular mechanisms governing cardiac development holds considerable promise for the development of innovative therapies for various cardiac ailments. While significant progress in uncovering novel transcriptional and epigenetic regulators of heart development has been made, the exploration of post-translational mechanisms that influence this process has lagged. Culling-RING E3 ubiquitin ligases (CRLs), the largest family of ubiquitin ligases, …

Augusta: From Rna-Seq To Gene Regulatory Networks And Boolean Models, Jana Musilova, Zdenek Vafek, Bhanwar Lal Puniya, Ralf Zimmer, Tomáš Helikar, Karel Sedlar

Department of Biochemistry: Faculty Publications

Computational models of gene regulations help to understand regulatory mechanisms and are extensively used in a wide range of areas, e.g., biotechnology or medicine, with significant benefits. Unfortunately, there are only a few computational gene regulatory models of whole genomes allowing static and dynamic analysis due to the lack of sophisticated tools for their reconstruction. Here, we describe Augusta, an open-source Python package for Gene Regulatory Network (GRN) and Boolean Network (BN) inference from the high-throughput gene expression data. Augusta can reconstruct genome-wide models suitable for static and dynamic analyses. Augusta uses a unique approach where the first estimation of …

The Multifunctional Catalytic Hemoglobin Fromamphitrite Ornata: Protocols On Isolation, Taxonomic Identification, Protein Extraction, Purification, And Characterization, Anna L. Husted, Lauren A. Presnar, R. Kevin Blackburn, Joseph L. Staton, Stephen A. Borgiani, Edward L. D'Antonio

Natural Sciences Faculty Publications

The multifunctional catalytic hemoglobin from the terebellid polychaete Amphitrite ornata, also named dehaloperoxidase (AoDHP), utilizes the typical oxygen transport function in addition to four observed activities involved in substrate oxidation. The multifunctional ability of AoDHP is presently a rare observation, and there exists a limitation for how novel dehaloperoxidases can be identified from macrobenthic infauna. In order to discover more infaunal DHP-bearing candidates, we have devised a facilitated method for an accurate taxonomic identification that places visual and molecular taxonomic approaches in parallel. Traditional visual taxonomic species identification by the non-specialist, at least for A. ornata or even for other …

Evaluating The Response Of Glycine Soja Accessions To Fungal Pathogen Macrophomina Phaseolina During Seedling Growth, Shirley Jacquet, Layla Rashad, Sonia Viera, Francisco Reta, Juan Reta

Biological Science Student Working Papers

Charcoal rot caused by the fungal pathogen Macrophomina phaseolina (Tassi) Goid is one of various devastating soybean (Glycine max (L.) Merr.) diseases, which can severely reduce crop yield. The investigation into the genetic potential for charcoal rot resistance of wild soybean (Glycine soja) accessions will enrich our understanding of the impact of soybean domestication on disease resistance; moreover, the identified charcoal rot-resistant lines can be used to improve soybean resistance to charcoal rot. The objective of this study was to evaluate the resistance of wild soybean accessions to M. phaseolina at the seedling stage and thereby select the disease-resistant lines. …

Changes In Lipid Profiles Of Epileptic Mouse Model, Alicia Johnson, Ryan A. Grove, Deepak Madhavan, Cory Honsinger Thomas Boone, Camila Pereira Braga, Hannah Kyllo, Kaeli Samson, Timothy Simeone, Tomáš Helikar, Corrine K. Hanson, Jiri Adamec

Department of Biochemistry: Faculty Publications

Introduction—Approximately 1% of the world’s population is impacted by epilepsy, a chronic neurological disorder characterized by seizures. One-third of epileptic patients are resistant to AEDs, or have medically refractory epilepsy (MRE). One non-invasive treatment that exists for MRE includes the ketogenic diet, a high-fat, low-carbohydrate diet. Despite the KD’s success in seizure attenuation, it has a few risks and its mechanisms remain poorly understood. The KD has been shown to improve metabolism and mitochondrial function in epileptic phenotypes. Potassium channels have implications in epileptic conditions as they have dual roles as metabolic sensors and control neuronal excitation.

Objectives—The …

Bola3 And Nfu1 Link Mitoribosome Iron–Sulfur Cluster Assembly To Multiple Mitochondrial Dysfunctions Syndrome, Hui Zhong, Alexandre Janer, Oleh Khalimonchuk, Hana Antonicka, Eric A. Shoubridge, Antoni Barrientos

Department of Biochemistry: Faculty Publications

The human mitochondrial ribosome contains three [2Fe–2S] clusters whose assembly pathway, role, and implications for mitochondrial and metabolic diseases are unknown. Here, structure-function correlation studies show that the clusters play a structural role during mitoribosome assembly. To uncover the assembly pathway, we have examined the effect of silencing the e xpression of Fe–S cluster biosynthetic and delivery factors on mitoribosome stability. We find that the mitoribosome receives its [²Fe–2S] clusters from the GLRX5-BOLA3 node. Additionally, the assembly of the small subunit depends on the mitoribosome biogenesis factor METTL17, recently reported containing a [⁴Fe–4S] cluster, which we …

Raman Spectral Cytopathology For Cancer Diagnostic Applications, Fiona Lyng, Damian Traynor, Isha Behl, Declan O'Dea, Hugh Byrne

Books/Book Chapters

Histopathology is currently regarded as the “gold standard” patient sampling technique for cancer diagnostics. It involves extraction of a tissue sample, or biopsy, which is then sectioned and stained for evaluation by a pathologist [1]. Cytopathology, on the other hand, is the field of disease diagnosis at the cellular level [2], necessitating only the microscopic evaluation of samples of cells which can be harvested by less invasive methods such as exfoliative brushing, needle probes or from biofluids. Based on the identification of abnormalities in cellular appearance, cytopathology is currently widely used to aid in the screening of cancer, as well …

Factors Influencing Instructors’ Adoption And Continued Use Of Computing Science Technologies: A Case Study In The Context Of Cell Collective, Changsoo Song, Resa M. Helikar, Wendy Smith, Tomáš Helikar

Department of Biochemistry: Faculty Publications

Acquiring computational modeling and simulation skills has become ever more critical for students in life sciences courses at the secondary and tertiary levels. Many modeling and simulation tools have been created to help instructors nurture those skills in their classrooms. Understanding the factors that may motivate instructors to use such tools is crucial to improve students’ learning, especially for having authentic modeling and simulation learning experiences. This study designed and tested a decomposed technology acceptance model in which the perceived usefulness and perceived ease of use constructs are split between the teaching and learning sides of the technology to examine …

Cobalt-Sulfur Coordination Chemistry Drives B12 Loading Onto Methionine Synthase, Romila Mascarenhas, Arkajit Guha, Zhu Li, Markus Ruetz, Sojin An, Javier Seravalli, Ruma Banerjee

Department of Biochemistry: Faculty Publications

Cobalt-sulfur (Co-S) coordination is labile to both oxidation and reduction chemistry and is rarely seen in Nature. Cobalamin (or vitamin B12) is an essential cobalt-containing organometallic cofactor in mammals, and is escorted via an intricate network of chaperones to a single cytoplasmic target, methionine synthase. In this study, we report that the human cobalamin trafficking protein, MMADHC, exploits the chemical lability of Co-S coordination, for cofactor off-loading onto methionine synthase. Cys-261 on MMADHC serves as the -axial ligand to cobalamin. Complex formation between MMADHC and methionine synthase is signaled by loss of the lower axial nitrogen ligand, leading to five-coordinate …

Editorial: Rising Stars In Microbial Physiology And Metabolism: 2022, Nicole R. Buan, Ulrike Kappler

Department of Biochemistry: Faculty Publications

This Research Topic was initiated to highlight work by young authors, the rising stars in the field of microbial physiology and metabolism. Microbial physiology and metabolism is an interdisciplinary field of research that seeks to uncover how the metabolic pathways of a cell work together to determine cell fate and function, whether that be growth, replication, pathogenicity, predation, respiration and fermentation, homeostasis or death. Ultimately, researchers like the ones featured here seek to integrate biological information and physicochemical parameters to try to find the underlying rules governing microbial function so that we can understand, predict and design microbes and microbial …

Pyrroline-5-Carboxylate Reductase-2 Promotes Colorectal Carcinogenesis By Modulating Microtubule-Associated Serine/Threonine Kinase-Like/Wnt/Β-Catenin Signaling, Raju Lama Tamang, Balawant Kumar, Sagar M. Patel, Ishwor Thapa, Alshomrani Ahmad, Vikas Kumar, Rizwan Ahmad, Donald F. Becker, Dundy Kiran Bastola, Punita Dhawan, Amar B. Singh

Department of Biochemistry: Faculty Publications

Background: Despite significant progress in clinical management, colorectal cancer (CRC) remains the third most common cause of cancer-related deaths. A positive association between PYCR2 (pyrroline-5-carboxylate reductase-2), a terminal enzyme of proline metabolism, and CRC aggressiveness was recently reported. However, how PYCR2 promotes colon carcinogenesis remains ill understood. Methods: A comprehensive analysis was performed using publicly available cancer databases and CRC patient cohorts. Proteomics and biochemical evaluations were performed along with genetic manipulations and in vivo tumor growth assays to gain a mechanistic understanding. Results: PYCR2 expression was significantly upregulated in CRC and associated with poor patient survival, specifically among PYCR …

Chloroquine And Cytosolic Galectins Affect Endosomal Escape Of Antisense Oligonucleotides After Stabilin-Mediated Endocytosis, Ekta Pandey, Ed Harris

Department of Biochemistry: Faculty Publications

Non-DNA-binding Stabilin-2/HARE receptors expressed on liver sinusoidal endothelial cells specifically bind to and internalize several classes of phosphorothioate antisense oligonucleotides (PS-ASOs). After Stabilin-mediated uptake, PS-ASOs are trafficked within endosomes (>97%–99%), ultimately resulting in destruction in the lysosome. The ASO entrapment in endosomes lowers therapeutic efficacy, thereby increasing the overall dose for patients. Here, we use confocal microscopy to characterize the intracellular route transverse by PS-ASOs after Stabilin receptor-mediated uptake in stable recombinant Stabilin-1 and -2 cell lines. We found that PS-ASOs as well as the Stabilin-2 receptor transverse the classic path: clathrincoated vesicle-early endosome-late endosome-lysosome. Chloroquine exposure facilitated endosomal …