Human trans-editing enzyme displays tRNA acceptor-stem specificity and relaxed amino acid selectivity [Enzymology]
Accurate translation of genetic information into proteins is vital for cell sustainability. ProXp-ala prevents proteome-wide Pro-to-Ala mutations by hydrolyzing misacylated Ala-tRNAPro, which is synthesized by prolyl-tRNA synthetase. Bacterial ProXp-ala was previously shown to combine a size-based exclusion mechanism with conformational and chemical selection for the recognition of the alanyl moiety, whereas tRNAPro is selected via recognition of tRNA acceptor-stem elements G72 and A73. The identity of these critical bases changed during evolution with eukaryotic cytosolic tRNAPro possessing a cytosine at the corresponding...
Source: Journal of Biological Chemistry - November 27, 2020 Category: Chemistry Authors: Oscar Vargas-Rodriguez, Marina Bakhtina, Daniel McGowan, Jawad Abid, Yuki Goto, Hiroaki Suga, Karin Musier-Forsyth Tags: Editors ' Picks Source Type: research
Increasing kinase domain proximity promotes MST2 autophosphorylation during Hippo signaling [Enzymology]
The Hippo pathway plays an important role in developmental biology, mediating organ size by controlling cell proliferation through the activity of a core kinase cassette. Multiple upstream events activate the pathway, but how each controls this core kinase cassette is not fully understood. Activation of the core kinase cassette begins with phosphorylation of the kinase MST1/2 (also known as STK3/4). Here, using a combination of in vitro biochemistry and cell-based assays, including chemically induced dimerization and single-molecule pulldown, we revealed that increasing the proximity of adjacent kinase domains, rather than...
Source: Journal of Biological Chemistry - November 20, 2020 Category: Chemistry Authors: Thao Tran, Jaba Mitra, Taekjip Ha, Jennifer M. Kavran Tags: Editors ' Picks Source Type: research
Template-dependent inhibition of coronavirus RNA-dependent RNA polymerase by remdesivir reveals a second mechanism of action [Enzymology]
Remdesivir (RDV) is a direct-acting antiviral agent that is used to treat patients with severe coronavirus disease 2019 (COVID-19). RDV targets the viral RNA-dependent RNA polymerase (RdRp) of severe acute respiratory syndrome coronavirus 2 (SARS–CoV-2). We have previously shown that incorporation of the active triphosphate form of RDV (RDV-TP) at position i causes delayed chain termination at position i + 3. Here we demonstrate that the S861G mutation in RdRp eliminates chain termination, which confirms the existence of a steric clash between Ser-861 and the incorporated RDV-TP. With WT RdRp, increasing concentrations o...
Source: Journal of Biological Chemistry - November 20, 2020 Category: Chemistry Authors: Egor P. Tchesnokov, Calvin J. Gordon, Emma Woolner, Dana Kocinkova, Jason K. Perry, Joy Y. Feng, Danielle P. Porter, Matthias Gotte Tags: Enzymology Source Type: research
Emerging roles of the MAGE protein family in stress response pathways [Protein Synthesis and Degradation]
The melanoma antigen (MAGE) proteins all contain a MAGE homology domain. MAGE genes are conserved in all eukaryotes and have expanded from a single gene in lower eukaryotes to ∼40 genes in humans and mice. Whereas some MAGEs are ubiquitously expressed in tissues, others are expressed in only germ cells with aberrant reactivation in multiple cancers. Much of the initial research on MAGEs focused on exploiting their antigenicity and restricted expression pattern to target them with cancer immunotherapy. Beyond their potential clinical application and role in tumorigenesis, recent studies have shown that MAGE proteins regul...
Source: Journal of Biological Chemistry - November 20, 2020 Category: Chemistry Authors: Rebecca R. Florke Gee, Helen Chen, Anna K. Lee, Christina A. Daly, Benjamin A. Wilander, Klementina Fon Tacer, Patrick Ryan Potts Tags: JBC Reviews Source Type: research
Overlapping function of Hrd1 and Ste24 in translocon quality control provides robust channel surveillance [Protein Synthesis and Degradation]
In this study, we found that two TQC enzymes, the ER-associated degradation ubiquitin ligase Hrd1 and zinc metalloprotease Ste24, promote degradation of characterized translocon-associated substrates of the other enzyme in Saccharomyces cerevisiae. Although both enzymes contribute to substrate turnover, our results suggest a prominent role for Hrd1 in TQC. Yeast lacking both Hrd1 and Ste24 exhibit a profound growth defect, consistent with overlapping function. Remarkably, two mutations that mildly perturb post-translational translocation and reduce the extent of aberrant translocon engagement by a model substrate diminish ...
Source: Journal of Biological Chemistry - November 20, 2020 Category: Chemistry Authors: Avery M. Runnebohm, Kyle A. Richards, Courtney Broshar Irelan, Samantha M. Turk, Halie E. Vitali, Christopher J. Indovina, Eric M. Rubenstein Tags: Accelerated Communications Source Type: research
A structural view of PA2G4 isoforms with opposing functions in cancer [Computational Biology]
The role of proliferation-associated protein 2G4 (PA2G4), alternatively known as ErbB3-binding protein 1 (EBP1), in cancer has become apparent over the past 20 years. PA2G4 expression levels are correlated with prognosis in a range of human cancers, including neuroblastoma, cervical, brain, breast, prostate, pancreatic, hepatocellular, and other tumors. There are two PA2G4 isoforms, PA2G4-p42 and PA2G4-p48, and although both isoforms of PA2G4 regulate cellular growth and differentiation, these isoforms often have opposing roles depending on the context. Therefore, PA2G4 can function either as a contextual tumor suppressor ...
Source: Journal of Biological Chemistry - November 20, 2020 Category: Chemistry Authors: Brendan W. Stevenson, Michael A. Gorman, Jessica Koach, Belamy B. Cheung, Glenn M. Marshall, Michael W. Parker, Jessica K. Holien Tags: JBC Reviews Source Type: research
Expression of a TMC6-TMC8-CIB1 heterotrimeric complex in lymphocytes is regulated by each of the components [Molecular Bases of Disease]
The TMC genes encode a set of homologous transmembrane proteins whose functions are not well understood. Biallelic mutations in either TMC6 or TMC8 are detected in more than half of cases of the pre-malignant skin disease epidermodysplasia verruciformis (EV). It is controversial whether EV induced by mutations in TMC6 or TMC8 originates from keratinocyte or lymphocyte defects. Quantification of TMC6 and TMC8 RNA levels in various organs revealed that lymphoid tissues have the highest levels of expression of both genes, and custom antibodies confirmed protein expression in mouse lymphocytes. To study the function of these p...
Source: Journal of Biological Chemistry - November 20, 2020 Category: Chemistry Authors: Chuan-Jin Wu, Xing Li, Connie L. Sommers, Kiyoto Kurima, Sunmee Huh, Grace Bugos, Lijin Dong, Wenmei Li, Andrew J. Griffith, Lawrence E. Samelson Tags: Signal Transduction Source Type: research
Uncoupling the hydrolysis of lipid-linked oligosaccharide from the oligosaccharyl transfer reaction by point mutations in yeast oligosaccharyltransferase [Enzymology]
Oligosaccharyltransferase (OST) is responsible for the first step in the N-linked glycosylation, transferring an oligosaccharide chain onto asparagine residues to create glycoproteins. In the absence of an acceptor asparagine, OST hydrolyzes the oligosaccharide donor, releasing free N-glycans (FNGs) into the lumen of the endoplasmic reticulum (ER). Here, we established a purification method for mutated OSTs using a high-affinity epitope tag attached to the catalytic subunit Stt3, from yeast cells co-expressing the WT OST to support growth. The purified OST protein with mutations is useful for wide-ranging biochemical exper...
Source: Journal of Biological Chemistry - November 20, 2020 Category: Chemistry Authors: Takahiro Yamasaki, Daisuke Kohda Tags: Glycobiology and Extracellular Matrices Source Type: research
E3 ubiquitin ligase PARK2, an inhibitor of melanoma cell growth, is repressed by the oncogenic ERK1/2-ELK1 transcriptional axis [Signal Transduction]
Malignant melanoma, the most aggressive form of skin cancer, is characterized by high prevalence of BRAF/NRAS mutations and hyperactivation of extracellular signal-regulated kinase 1 and 2 (ERK1/2), mitogen-activated protein kinases (MAPK), leading to uncontrolled melanoma growth. Efficacy of current targeted therapies against mutant BRAF or MEK1/2 have been hindered by existence of innate or development of acquired resistance. Therefore, a better understanding of the mechanisms controlled by MAPK pathway driving melanogenesis will help develop new treatment approaches targeting this oncogenic cascade. Here, we identify E3...
Source: Journal of Biological Chemistry - November 20, 2020 Category: Chemistry Authors: Valentina Montagnani, Luisa Maresca, Alessandro Apollo, Sara Pepe, Ryan M. Carr, Martin E. Fernandez-Zapico, Barbara Stecca Tags: Molecular Bases of Disease Source Type: research
Allosteric inhibition of MTHFR prevents futile SAM cycling and maintains nucleotide pools in one-carbon metabolism [Enzymology]
Methylenetetrahydrofolate reductase (MTHFR) links the folate cycle to the methionine cycle in one-carbon metabolism. The enzyme is known to be allosterically inhibited by SAM for decades, but the importance of this regulatory control to one-carbon metabolism has never been adequately understood. To shed light on this issue, we exchanged selected amino acid residues in a highly conserved stretch within the regulatory region of yeast MTHFR to create a series of feedback-insensitive, deregulated mutants. These were exploited to investigate the impact of defective allosteric regulation on one-carbon metabolism. We observed a s...
Source: Journal of Biological Chemistry - November 20, 2020 Category: Chemistry Authors: Muskan Bhatia, Jyotika Thakur, Shradha Suyal, Ruchika Oniel, Rahul Chakraborty, Shalini Pradhan, Monika Sharma, Shantanu Sengupta, Sunil Laxman, Shyam Kumar Masakapalli, Anand Kumar Bachhawat Tags: Metabolism Source Type: research
Decrease of core 2 O-glycans on synovial lubricin in osteoarthritis reduces galectin-3 mediated crosslinking [Molecular Bases of Disease]
The synovial fluid glycoprotein lubricin (also known as proteoglycan 4) is a mucin-type O-linked glycosylated biological lubricant implicated to be involved in osteoarthritis (OA) development. Lubricin's ability to reduce friction is related to its glycosylation consisting of sialylated and unsialylated Tn-antigens and core 1 and core 2 structures. The glycans on lubricin have also been suggested to be involved in crosslinking and stabilization of the lubricating superficial layer of cartilage by mediating interaction between lubricin and galectin-3. However, with the spectrum of glycans being found on lubricin, the glycan...
Source: Journal of Biological Chemistry - November 20, 2020 Category: Chemistry Authors: Sarah A. Flowers, Kristina A. Thomsson, Liaqat Ali, Shan Huang, Yolanda Mthembu, Suresh C. Regmi, Jan Holgersson, Tannin A. Schmidt, Ola Rolfson, Lena I. Bȷorkman, Martina Sundqvist, Anna Karlsson–Bengtsson, Gregory D. Jay, Thomas Eisler, Rom Tags: Glycobiology and Extracellular Matrices Source Type: research
Substrate binding tunes the reactivity of hispidin 3-hydroxylase, a flavoprotein monooxygenase involved in fungal bioluminescence [Enzymology]
Fungal bioluminescence was recently shown to depend on a unique oxygen-dependent system of several enzymes. However, the identities of the enzymes did not reveal the full biochemical details of this process, as the enzymes do not bear resemblance to those of other luminescence systems, and thus the properties of the enzymes involved in this fascinating process are still unknown. Here, we describe the characterization of the penultimate enzyme in the pathway, hispidin 3-hydroxylase, from the luminescent fungus Mycena chlorophos (McH3H), which catalyzes the conversion of hispidin to 3-hydroxyhispidin. 3-Hydroxyhispidin acts ...
Source: Journal of Biological Chemistry - November 20, 2020 Category: Chemistry Authors: Yapei Tong, Milos Trajkovic, Simone Savino, Willem J. H. van Berkel, Marco W. Fraaije Tags: Enzymology Source Type: research
Proteolytic cleavage of Podocin by Matriptase exacerbates podocyte injury [Cell Biology]
Podocyte injury is a critical step toward the progression of renal disease and is often associated with a loss of slit diaphragm proteins, including Podocin. Although there is a possibility that the extracellular domain of these slit diaphragm proteins can be a target for a pathological proteolysis, the precise mechanism driving the phenomenon remains unknown. Here we show that Matriptase, a membrane-anchored protein, was activated at podocytes in CKD patients and mice, whereas Matriptase inhibitors slowed the progression of mouse kidney disease. The mechanism could be accounted for by an imbalance favoring Matriptase over...
Source: Journal of Biological Chemistry - November 20, 2020 Category: Chemistry Authors: Shota Ozawa, Masaya Matsubayashi, Hitoki Nanaura, Motoko Yanagita, Kiyoshi Mori, Katsuhiko Asanuma, Nobuyuki Kajiwara, Kazuyuki Hayashi, Hiroshi Ohashi, Masato Kasahara, Hideki Yokoi, Hiroaki Kataoka, Eiichiro Mori, Takahiko Nakagawa Tags: Molecular Bases of Disease Source Type: research
GGPP depletion initiates metaflammation through disequilibrating CYB5R3-dependent eicosanoid metabolism [Metabolism]
Metaflammation is a primary inflammatory complication of metabolic disorders characterized by altered production of many inflammatory cytokines, adipokines, and lipid mediators. Whereas multiple inflammation networks have been identified, the mechanisms by which metaflammation is initiated have long been controversial. As the mevalonate pathway (MVA) produces abundant bioactive isoprenoids and abnormal MVA has a phenotypic association with inflammation/immunity, we speculate that isoprenoids from the MVA may provide a causal link between metaflammation and metabolic disorders. Using a line with the MVA isoprenoid producer ...
Source: Journal of Biological Chemistry - November 20, 2020 Category: Chemistry Authors: Lisha Wei, Yan-Yan Zheng, Jie Sun, Pei Wang, Tao Tao, Yeqiong Li, Xin Chen, Yongjuan Sang, Danyang Chong, Wei Zhao, Yuwei Zhou, Ye Wang, Zhihui Jiang, Tiantian Qiu, Chao-Jun Li, Min-Sheng Zhu, Xuena Zhang Tags: Cell Biology Source Type: research
Histo-blood group antigens of glycosphingolipids predict susceptibility of human intestinal enteroids to norovirus infection [Microbiology]
The molecular mechanisms behind infection and propagation of human restricted pathogens such as human norovirus (HuNoV) have defied interrogation because they were previously unculturable. However, human intestinal enteroids (HIEs) have emerged to offer unique ex vivo models for targeted studies of intestinal biology, including inflammatory and infectious diseases. Carbohydrate-dependent histo-blood group antigens (HBGAs) are known to be critical for clinical infection. To explore whether HBGAs of glycosphingolipids contribute to HuNoV infection, we obtained HIE cultures established from stem cells isolated from jejunal bi...
Source: Journal of Biological Chemistry - November 20, 2020 Category: Chemistry Authors: Inga Rimkute, Konrad Thorsteinsson, Marcus Henricsson, Victoria R. Tenge, Xiaoming Yu, Shih–Ching Lin, Kei Haga, Robert L. Atmar, Nils Lycke, Jonas Nilsson, Mary K. Estes, Marta Bally, Goran Larson Tags: Glycobiology and Extracellular Matrices Source Type: research
