Comparison of type 5d autotransporter phospholipases demonstrates a correlation between high activity and intracellular pathogenic lifestyle
Autotransporters, or type 5 secretion systems, are widespread surface proteins of Gram-negative bacteria often associated with virulence functions. Autotransporters consist of an outer membrane β-barrel domain and an exported passenger. In the poorly studied type 5d subclass, the passenger is a patatin-like lipase. The prototype of this secretion pathway is PlpD of Pseudomonas aeruginosa, an opportunistic human pathogen. The PlpD passenger is a homodimer with phospholipase A1 (PLA1) activity. Based on sequencing data, PlpD-like proteins are present in many bacterial species. We characterized the enzymatic activity, sp...
Source: Biochemical Journal - September 24, 2019 Category: Biochemistry Authors: Trunk, T., Casasanta, M. A., Yoo, C. C., Slade, D. J., Leo, J. C. Tags: Research Articles Source Type: research

The sugar code: letters and vocabulary, writers, editors and readers and biosignificance of functional glycan-lectin pairing
Ubiquitous occurrence in Nature, abundant presence at strategically important places such as the cell surface and dynamic shifts in their profile by diverse molecular switches qualifies the glycans to serve as versatile biochemical signals. However, their exceptional structural complexity often prevents one noting how simple the rules of objective-driven assembly of glycan-encoded messages are. This review is intended to provide a tutorial for a broad readership. The principles of why carbohydrates meet all demands to be the coding section of an information transfer system, and this at unsurpassed high density, are explain...
Source: Biochemical Journal - September 24, 2019 Category: Biochemistry Authors: Kaltner, H., Abad-Rodriguez, J., Corfield, A. P., Kopitz, J., Gabius, H.-J. Tags: Review Articles Source Type: research

Conserved bases for the initial cyclase in gibberellin biosynthesis: from bacteria to plants
All land plants contain at least one class II diterpene cyclase (DTC), which utilize an acid-base catalytic mechanism, for the requisite production of ent-copalyl diphosphate (ent-CPP) in gibberellin A (GA) phytohormone biosynthesis. These ent-CPP synthases (CPSs) are hypothesized to be derived from ancient bacterial origins and, in turn, to have given rise to the frequently observed additional DTCs utilized in more specialized plant metabolism. However, such gene duplication and neo-functionalization has occurred repeatedly, reducing the utility of phylogenetic analyses. Support for evolutionary scenarios can be found in ...
Source: Biochemical Journal - September 24, 2019 Category: Biochemistry Authors: Lemke, C., Potter, K. C., Schulte, S., Peters, R. J. Tags: Research Articles Source Type: research

Rubisco activation by wheat Rubisco activase isoform 2{beta} is insensitive to inhibition by ADP
Rubisco activase (Rca) is a catalytic chaperone that remodels the active site, promotes the release of inhibitors and restores catalytic competence to Rubisco. Rca activity and its consequent effect on Rubisco activation and photosynthesis are modulated by changes to the chloroplast environment induced by fluctuations in light levels that reach the leaf, including redox status and adenosine diphosphate (ADP)/adenosine triphosphate (ATP) ratio. The Triticum aestivum (wheat) genome encodes for three Rca protein isoforms: 1β (42.7 kDa), 2β (42.2 kDa) and 2α (46.0 kDa). The regulatory properties ...
Source: Biochemical Journal - September 24, 2019 Category: Biochemistry Authors: Perdomo, J. A., Degen, G. E., Worrall, D., Carmo-Silva, E. Tags: Research Articles Source Type: research

A polyamine-independent role for S-adenosylmethionine decarboxylase
The only known function of S-adenosylmethionine decarboxylase (AdoMetDC) is to supply, with its partner aminopropyltransferase enzymes such as spermidine synthase (SpdSyn), the aminopropyl donor for polyamine biosynthesis. Polyamine spermidine is probably essential for the growth of all eukaryotes, most archaea and many bacteria. Two classes of AdoMetDC exist, the prokaryotic class 1a and 1b forms, and the eukaryotic class 2 enzyme, which is derived from an ancient fusion of two prokaryotic class 1b genes. Herein, we show that ‘eukaryotic' class 2 AdoMetDCs are found in bacteria and are enzymatically functional. Howe...
Source: Biochemical Journal - September 20, 2019 Category: Biochemistry Authors: Li, B., Kurihara, S., Kim, S. H., Liang, J., Michael, A. J. Tags: Research Articles Source Type: research

Ufd1 phosphorylation at serine 229 negatively regulates endoplasmic reticulum-associated degradation by inhibiting the interaction of Ufd1 with VCP
Misfolded proteins in the endoplasmic reticulum (ER) are removed through multistep processes termed ER-associated degradation (ERAD). Valosin-containing protein (VCP) plays a crucial role in ERAD as the interaction of ubiquitin fusion degradation protein 1 (Ufd1) with VCP via its SHP box motif (228F-S-G-S-G-N-R-L235) is required for ERAD. However, the mechanisms by which the VCP–Ufd1 interaction is regulated are not well understood. Here, we found that the serine 229 residue located in the Ufd1 SHP box is phosphorylated in vitro and in vivo by cyclic adenosine monophosphate-dependent protein kinase A (PKA), with this...
Source: Biochemical Journal - September 20, 2019 Category: Biochemistry Authors: Nguyen, Q.-A. T., Choi, J., Yang, J. K., Lee, S. Y. Tags: Research Articles Source Type: research

Valosin-containing protein mediates the ERAD of squalene monooxygenase and its cholesterol-responsive degron
Squalene monooxygenase (SM) is an essential rate-limiting enzyme in cholesterol synthesis. SM degradation is accelerated by excess cholesterol, and this requires the first 100 amino acids of SM (SM N100). This process is part of a protein quality control pathway called endoplasmic reticulum-associated degradation (ERAD). In ERAD, SM is ubiquitinated by MARCH6, an E3 ubiquitin ligase located in the endoplasmic reticulum (ER). However, several details of the ERAD process for SM remain elusive, such as the extraction mechanism from the ER membrane. Here, we used SM N100 fused to GFP (SM N100-GFP) as a model degron to investig...
Source: Biochemical Journal - September 20, 2019 Category: Biochemistry Authors: Chua, N. K., Scott, N. A., Brown, A. J. Tags: Research Articles Source Type: research

Structural and functional characterisation of human RNA helicase DHX8 provides insights into the mechanism of RNA-stimulated ADP release
This study provides an in-depth understanding of the activity of DHX8 and contributes insights into the RNA-unwinding mechanisms of the DEAH-box helicase family. (Source: Biochemical Journal)
Source: Biochemical Journal - September 13, 2019 Category: Biochemistry Authors: Felisberto-Rodrigues, C., Thomas, J. C., McAndrew, C., Le Bihan, Y.-V., Burke, R., Workman, P., van Montfort, R. L. M. Tags: Research Articles Source Type: research

Correction: Tissue-specific characterization of mitochondrial branched-chain keto acid oxidation using a multiplexed assay platform
(Source: Biochemical Journal)
Source: Biochemical Journal - September 13, 2019 Category: Biochemistry Authors: Goldberg, E. J., Buddo, K. A., McLaughlin, K. L., Fernandez, R. F., Pereyra, A. S., Psaltis, C. E., Lin, C.-T., Hagen, J. T., Boykov, I. N., Nguyen, T. K., Gowdy, K. M., Ellis, J. M., Neufer, P. D., McClung, J. M., Fisher-Wellman, K. H. Tags: Corrections Source Type: research

Correction: Designing active RNF4 monomers by introducing a tryptophan: avidity towards E2~Ub conjugates dictates the activity of ubiquitin RING E3 ligases
(Source: Biochemical Journal)
Source: Biochemical Journal - September 13, 2019 Category: Biochemistry Authors: Sarkar, S., Behera, A. P., Borar, P., Banka, P. A., Datta, A. B. Tags: Corrections Source Type: research

Correction: Structural insights into the nanomolar affinity of RING E3 ligase ZNRF1 for Ube2N and its functional implications
(Source: Biochemical Journal)
Source: Biochemical Journal - September 13, 2019 Category: Biochemistry Authors: Behera, A. P., Naskar, P., Agarwal, S., Banka, P. A., Poddar, A., Datta, A. B. Tags: Corrections Source Type: research

RhoBTB1 interacts with ROCKs and inhibits invasion
We report that RhoBTB1 depletion increases prostate cancer cell invasion and induces elongation in Matrigel, a phenotype similar to that induced by depletion of ROCK1 and ROCK2. We demonstrate that RhoBTB1 associates with ROCK1 and ROCK2 and its association with ROCK1 is via its Rho domain. The Rho domain binds to the coiled-coil region of ROCK1 close to its kinase domain. We identify two amino acids within the Rho domain that alter RhoBTB1 association with ROCK1. RhoBTB1 is a substrate for ROCK1, and mutation of putative phosphorylation sites reduces its association with Cullin3, a scaffold for ubiquitin ligases. We propo...
Source: Biochemical Journal - September 13, 2019 Category: Biochemistry Authors: Haga, R. B., Garg, R., Collu, F., Borda D'Agua, B., Menendez, S. T., Colomba, A., Fraternali, F., Ridley, A. J. Tags: Research Articles Source Type: research

Role and regulation of class-C flavodiiron proteins in photosynthetic organisms
The regulation of photosynthesis is crucial to efficiently support the assimilation of carbon dioxide and to prevent photodamage. One key regulatory mechanism is the pseudo-cyclic electron flow (PCEF) mediated by class-C flavodiiron proteins (FLVs). These enzymes use electrons coming from Photosystem I (PSI) to reduce oxygen to water, preventing over-reduction in the acceptor side of PSI. FLVs are widely distributed among organisms performing oxygenic photosynthesis and they have been shown to be fundamental in many different conditions such as fluctuating light, sulfur deprivation and plant submersion. Moreover, since FLV...
Source: Biochemical Journal - September 13, 2019 Category: Biochemistry Authors: Alboresi, A., Storti, M., Cendron, L., Morosinotto, T. Tags: Review Articles Source Type: research

Mitofusins modulate the increase in mitochondrial length, bioenergetics and secretory phenotype in therapy-induced senescent melanoma cells
Cellular senescence is an endpoint of chemotherapy, and targeted therapies in melanoma and the senescence-associated secretory phenotype (SASP) can affect tumor growth and microenvironment, influencing treatment outcomes. Metabolic interventions can modulate the SASP, and an enhanced mitochondrial energy metabolism supports resistance to therapy in melanoma cells. Herein, we assessed the mitochondrial function of therapy-induced senescent melanoma cells obtained after exposing the cells to temozolomide (TMZ), a methylating chemotherapeutic agent. Senescence induction in melanoma was accompanied by a substantial increase in...
Source: Biochemical Journal - September 10, 2019 Category: Biochemistry Authors: Martinez, J., Tarallo, D., Martinez-Palma, L., Victoria, S., Bresque, M., Rodriguez-Bottero, S., Marmisolle, I., Escande, C., Cassina, P., Casanova, G., Bollati-Fogolin, M., Agorio, C., Moreno, M., Quijano, C. Tags: Research Articles Source Type: research

Interactions between motor domains in kinesin-14 Ncd -- a molecular dynamics study
Minus-end directed, non-processive kinesin-14 Ncd is a dimeric protein with C-terminally located motor domains (heads). Generation of the power-stroke by Ncd consists of a lever-like rotation of a long superhelical ‘stalk’ segment while one of the kinesin's heads is bound to the microtubule. The last ~30 amino acids of Ncd head play a crucial but still poorly understood role in this process. Here, we used accelerated molecular dynamics simulations to explore the conformational dynamics of several systems built upon two crystal structures of Ncd, the asymmetrical T436S mutant in pre-stroke/post-stroke conformati...
Source: Biochemical Journal - September 10, 2019 Category: Biochemistry Authors: Ludwiczak, J., Szczesna, E., da Silva Neto, A. M., Cieplak, P., Kasprzak, A. A., Jarmuła, A. Tags: Research Articles Source Type: research

The synthesis of branched-chain fatty acids is limited by enzymatic decarboxylation of ethyl- and methylmalonyl-CoA
Most fatty acids (FAs) are straight chains and are synthesized by fatty acid synthase (FASN) using acetyl-CoA and malonyl-CoA units. Yet, FASN is known to be promiscuous as it may use methylmalonyl-CoA instead of malonyl-CoA and thereby introduce methyl-branches. We have recently found that the cytosolic enzyme ECHDC1 degrades ethylmalonyl-CoA and methylmalonyl-CoA, which presumably result from promiscuous reactions catalyzed by acetyl-CoA carboxylase on butyryl- and propionyl-CoA. Here, we tested the hypothesis that ECHDC1 is a metabolite repair enzyme that serves to prevent the formation of methyl- or ethyl-branched FAs ...
Source: Biochemical Journal - August 30, 2019 Category: Biochemistry Authors: Dewulf, J. P., Gerin, I., Rider, M. H., Veiga-da-Cunha, M., Van Schaftingen, E., Bommer, G. T. Tags: Research Articles Source Type: research

Broken force dispersal network in tip-links by the mutations at the Ca2+-binding residues induces hearing-loss
Tip-link as force-sensor in hearing conveys the mechanical force originating from sound to ion-channels while maintaining the integrity of the entire sensory assembly in the inner ear. This delicate balance between structure and function of tip-links is regulated by Ca2+-ions present in endolymph. Mutations at the Ca2+-binding sites of tip-links often lead to congenital deafness, sometimes syndromic defects impairing vision along with hearing. Although such mutations are already identified, it is still not clear how the mutants alter the structure-function properties of the force-sensors associated with diseases. With an a...
Source: Biochemical Journal - August 30, 2019 Category: Biochemistry Authors: Hazra, J. P., Sagar, A., Arora, N., Deb, D., Kaur, S., Rakshit, S. Tags: Research Articles Source Type: research

Rhizobacteria AK1 remediates the toxic effects of salinity stress via regulation of endogenous phytohormones and gene expression in soybean
In conclusion, AK1 can mitigate salinity stress, increase plant growth and could be utilized as an eco-friendly bio-fertilizer under salinity stress. (Source: Biochemical Journal)
Source: Biochemical Journal - August 30, 2019 Category: Biochemistry Authors: Khan, M. A., Asaf, S., Khan, A. L., Jan, R., Kang, S.-M., Kim, K.-M., Lee, I.-J. Tags: Research Articles Source Type: research

A novel small molecule A2A adenosine receptor agonist, indirubin-3'-monoxime, alleviates lipid-induced inflammation and insulin resistance in 3T3-L1 adipocytes
Saturated free fatty acid-induced adipocyte inflammation plays a pivotal role in implementing insulin resistance and type 2 diabetes. Recent reports suggest A2A adenosine receptor (A2AAR) could be an attractive choice to counteract adipocyte inflammation and insulin resistance. Thus, an effective A2AAR agonist devoid of any toxicity is highly appealing. Here, we report that indirubin-3'-monoxime (I3M), a derivative of the bisindole alkaloid indirubin, efficiently binds and activates A2AAR which leads to the attenuation of lipid-induced adipocyte inflammation and insulin resistance. Using a combination of in silico virtual ...
Source: Biochemical Journal - August 30, 2019 Category: Biochemistry Authors: Choudhary, S. A., Bora, N., Banerjee, D., Arora, L., Das, A. S., Yadav, R., Klotz, K.-N., Pal, D., Jha, A. N., Dasgupta, S. Tags: Research Articles Source Type: research

Autoactivation and calpain-1-mediated shedding of hepsin in human hepatoma cells
In this study, we conducted site-directed mutagenesis, cell expression, plasma membrane protein labeling, trypsin digestion, Western blotting, and flow cytometry experiments in human hepatoma HepG2 cells, where hepsin was originally discovered, and SMMC-7721 cells. Our results show that hepsin is activated by autocatalysis on the cell surface but not intracellularly. Moreover, we show that hepsin undergoes ectodomain shedding. In the conditioned medium from HepG2 and SMMC-7721 cells, we detected a soluble fragment comprising nearly the entire extracellular region of hepsin. By testing protease inhibitors, gene knockdown, a...
Source: Biochemical Journal - August 28, 2019 Category: Biochemistry Authors: Wang, L., Zhang, C., Sun, S., Chen, Y., Hu, Y., Wang, H., Liu, M., Dong, N., Wu, Q. Tags: Research Articles Source Type: research

The PHD finger of Spp1 mediates histone modification cross-talk
Binding of the Spp1 PHD finger to histone H3K4me3 is sensitive to adjacent post-translational modifications in the histone tail. This commentary discusses the findings of He and colleagues [Biochem. J. 476, 1957–1973] which show that the PHD finger binds to H3K4me3 in a selective manner which is conserved in the Saccharomyces pombe and mammalian orthologues of Spp1. (Source: Biochemical Journal)
Source: Biochemical Journal - August 28, 2019 Category: Biochemistry Authors: Musselman, C. A., Kutateladze, T. G. Tags: Commentaries Source Type: research

Pic1, counteracting plant immunity signalling
Plants are equipped with versatile pattern recognition receptors (PRRs), which monitor their external environment and elicit defensive measures upon detection of potential risk for disease. Inside the cell, receptor-like cytoplasmic kinases (RLCKs) are key components of PRR signalling, but their molecular functions and regulatory interactions are not yet fully understood. In tomato, two RLCKs, Pti1a and Pti1b, are important signalling components that relay early defence signals elicited by bacterial flagellin, a conserved pattern common to various pathogenic and non-pathogenic microbes. An important question to resolve is ...
Source: Biochemical Journal - August 28, 2019 Category: Biochemistry Authors: Kangasjärvi, S. Tags: Commentaries Source Type: research

The Great Escape: how phosphatidylinositol 4-kinases and PI4P promote vesicle exit from the Golgi (and drive cancer)
Phosphatidylinositol 4-phosphate (PI4P) is a membrane glycerophospholipid and a major regulator of the characteristic appearance of the Golgi complex as well as its vesicular trafficking, signalling and metabolic functions. Phosphatidylinositol 4-kinases, and in particular the PI4KIIIβ isoform, act in concert with PI4P to recruit macromolecular complexes to initiate the biogenesis of trafficking vesicles for several Golgi exit routes. Dysregulation of Golgi PI4P metabolism and the PI4P protein interactome features in many cancers and is often associated with tumour progression and a poor prognosis. Increased expressio...
Source: Biochemical Journal - August 28, 2019 Category: Biochemistry Authors: Waugh, M. G. Tags: Review Articles Source Type: research

Crystal structures of plant inorganic pyrophosphatase, an enzyme with a moonlighting autoproteolytic activity
Inorganic pyrophosphatases (PPases, EC 3.6.1.1), which hydrolyze inorganic pyrophosphate to phosphate in the presence of divalent metal cations, play a key role in maintaining phosphorus homeostasis in cells. DNA coding inorganic pyrophosphatases from Arabidopsis thaliana (AtPPA1) and Medicago truncatula (MtPPA1) were cloned into a bacterial expression vector and the proteins were produced in Escherichia coli cells and crystallized. In terms of their subunit fold, AtPPA1 and MtPPA1 are reminiscent of other members of Family I soluble pyrophosphatases from bacteria and yeast. Like their bacterial orthologs, both plant PPase...
Source: Biochemical Journal - August 22, 2019 Category: Biochemistry Authors: Grzechowiak, M., Ruszkowski, M., Sliwiak, J., Szpotkowski, K., Sikorski, M., Jaskolski, M. Tags: Research Articles Source Type: research

The human transmembrane mucin MUC17 responds to TNF{alpha} by increased presentation at the plasma membrane
Transmembrane mucin MUC17 is an integral part of the glycocalyx as it covers the brush border membrane of small intestinal enterocytes and presents an extended O-glycosylated mucin domain to the intestinal lumen. Here, we identified two unknown phosphorylated serine residues, S4428 and S4492, in the cytoplasmic tail of human MUC17. We have previously demonstrated that MUC17 is anchored to the apical membrane domain via an interaction with the scaffolding protein PDZK1. S4492, localized in the C-terminal PDZ binding motif of MUC17, was mutated to generate phosphomimetic and phosphodeficient variants of MUC17. Using Caco-2 c...
Source: Biochemical Journal - August 22, 2019 Category: Biochemistry Authors: Schneider, H., Berger, E., Dolan, B., Martinez-Abad, B., Arike, L., Pelaseyed, T., Hansson, G. C. Tags: Research Articles Source Type: research

A gene cluster for taurine sulfur assimilation in an anaerobic human gut bacterium
Aminoethylsulfonate (taurine) is widespread in the environment and highly abundant in the human body. Taurine and other aliphatic sulfonates serve as sulfur sources for diverse aerobic bacteria, which carry out cleavage of the inert sulfonate C–S bond through various O2-dependent mechanisms. Taurine also serves as a sulfur source for certain strict anaerobic fermenting bacteria. However, the mechanism of C–S cleavage by these bacteria has long been a mystery. Here we report the biochemical characterization of an anaerobic pathway for taurine sulfur assimilation in a strain of Clostridium butyricum from the huma...
Source: Biochemical Journal - August 15, 2019 Category: Biochemistry Authors: Xing, M., Wei, Y., Hua, G., Li, M., Nanjaraj Urs, A. N., Wang, F., Hu, Y., Zhai, W., Liu, Y., Ang, E. L., Zhao, H., Zhang, Y. Tags: Research Articles Source Type: research

Cancerous inhibitor of protein phosphatase 2A (CIP2A) modifies energy metabolism via 5' AMP-activated protein kinase signalling in malignant cells
Cancerous inhibitor of protein phosphatase 2A (CIP2A) is an adverse biomarker across many malignancies. Using K562 cells engineered to have high or low CIP2A expression, we show that high CIP2A levels significantly bias cellular energy production towards oxidative phosphorylation (OXPHOS) rather than glycolysis. Mass spectrometric analysis of CIP2A interactors and isobaric tagging for relative and absolute protein quantitation (ITRAQ) experiments identified many associated proteins, several of which co-vary with CIP2A level. Many of these CIP2A associating and co-varying proteins are involved in energy metabolism including...
Source: Biochemical Journal - August 15, 2019 Category: Biochemistry Authors: Austin, J. A., Jenkins, R. E., Austin, G. M., Glenn, M. A., Dunn, K., Scott, L., Lucas, C. M., Clark, R. E. Tags: Research Articles Source Type: research

ELP3 Acetyltransferase is phosphorylated and regulated by the oncogenic anaplastic lymphoma kinase (ALK)
Protein lysine acetylation is one of the major posttranslational modifications (PTMs) with several thousands of proteins identified to be acetylated in mammalian tissues. Mechanistic studies have revealed important functions of acetylation in the regulation of protein function. Much less is known on how the acetyltransferases themselves are regulated. In the current study, we discover that the Elongator protein 3 (ELP3) acetyltransferase is modified by tyrosine phosphorylation. We demonstrate that the anaplastic lymphoma kinase (ALK) is the major tyrosine kinase responsible for ELP3 tyrosine phosphorylation. ELP3 is phosph...
Source: Biochemical Journal - August 15, 2019 Category: Biochemistry Authors: Li, M.-T., Liang, J.-Y., Sun, Y.-P., Jin, J., Xiong, Y., Guan, K.-L., Yuan, H.-X. Tags: Research Articles Source Type: research

New tools for an old question: dependence of ATP and bicarbonate for branched-chain keto acids oxidation
Branched-chain keto acids (BCKA) metabolism involves several well-regulated steps within mitochondria, requires cofactors, and is modulated according to the metabolic status of the cells. This regulation has made it challenging to utilize in vitro approaches to determine the contribution of branched-chain amino acid oxidation to energy production. These methodological issues were elegantly addressed in a recent publication within the Biochemical Journal. In this issue, Goldberg et al. [Biochem. J. (2019) 476, 1521–1537] demonstrated in a well-designed system the dependence of ATP and bicarbonate for BCKA full oxidati...
Source: Biochemical Journal - August 15, 2019 Category: Biochemistry Authors: Brunetta, H. S., Holloway, G. P. Tags: Commentaries Source Type: research

Simple rules govern the diversity of bacterial nicotianamine-like metallophores
In metal-scarce environments, some pathogenic bacteria produce opine-type metallophores mainly to face the host's nutritional immunity. This is the case of staphylopine, pseudopaline and yersinopine, identified in Staphylococcus aureus, Pseudomonas aeruginosa and Yersinia pestis, respectively. Depending on the species, these metallophores are synthesized by two (CntLM) or three enzymes (CntKLM), CntM catalyzing the last step of biosynthesis using diverse substrates (pyruvate or α-ketoglutarate), pathway intermediates (xNA or yNA) and cofactors (NADH or NADPH). Here, we explored the substrate specificity of CntM by co...
Source: Biochemical Journal - August 9, 2019 Category: Biochemistry Authors: Laffont, C., Brutesco, C., Hajjar, C., Cullia, G., Fanelli, R., Ouerdane, L., Cavelier, F., Arnoux, P. Tags: Research Articles Source Type: research

Discovery of novel inhibitors of ribosome biogenesis by innovative high throughput screening strategies
In this study, two high-throughput screens were used to identify ribosome biogenesis inhibitors. Our primary screen made use of the HaloTag selective labeling strategy to identify compounds that decreased the abundance of newly synthesized ribosomes in A375 malignant melanoma cells. This screen identified 5786 hit compounds. A subset of those initial hit compounds were tested using a secondary screen that directly measured pre-ribosomal RNA (pre-rRNA) abundance as a reporter of rRNA synthesis rate, using quantitative RT-PCR. From the secondary screen, we identified two structurally related compounds that are potent inhibit...
Source: Biochemical Journal - August 9, 2019 Category: Biochemistry Authors: Scull, C. E., Zhang, Y., Tower, N., Rasmussen, L., Padmalayam, I., Hunter, R., Zhai, L., Bostwick, R., Schneider, D. A. Tags: Research Articles Source Type: research

Gelatinase B/matrix metalloproteinase-9 and other neutrophil proteases switch off interleukin-2 activity
Interleukin 2 (IL-2) is critical for T cell development and homeostasis, being a key regulator of adaptive immune responses in autoimmunity, hypersensitivity reactions and cancer. Therefore, its abundance in serum and peripheral tissues needs tight control. Here, we described a new mechanism contributing to the immunobiology of IL-2. We demonstrated, both in biochemical and cell-based assays, that IL-2 is subject to proteolytic processing by neutrophil matrix metalloproteinase-9 (MMP-9). IL-2 fragments produced after cleavage by MMP-9 remained linked by a disulfide bond and displayed a reduced affinity for all IL-2 recepto...
Source: Biochemical Journal - August 9, 2019 Category: Biochemistry Authors: Rybakin, V., Stas, M., Ugarte-Berzal, E., Noppen, S., Vandooren, J., Van Aelst, I., Liekens, S., Proost, P., Opdenakker, G. Tags: Research Articles Source Type: research

Dissecting the cytochrome c2-reaction centre interaction in bacterial photosynthesis using single molecule force spectroscopy
We examined the interaction of two components of bacterial photosynthesis, cytochrome c2 and the reaction centre (RC) complex, using dynamic force spectroscopy and PeakForce quantitative nanomechanical imaging. RC–LH1–PufX complexes, attached to silicon nitride AFM probes and maintained in a photo-oxidised state, were lowered onto a silicon oxide substrate bearing dispersed, immobilised and reduced cytochrome c2 molecules. Microscale patterns of cytochrome c2 and the cyan fluorescent protein were used to validate the specificity of recognition between tip-attached RCs and surface-tethered cytochrome c2. Followi...
Source: Biochemical Journal - August 9, 2019 Category: Biochemistry Authors: Vasilev, C., Mayneord, G. E., Brindley, A. A., Johnson, M. P., Hunter, C. N. Tags: Research Articles Source Type: research

A biochemical comparison of fungal GH6 cellobiohydrolases
Cellobiohydrolases (CBHs) from glycoside hydrolase family 6 (GH6) make up an important part of the secretome in many cellulolytic fungi. They are also of technical interest, particularly because they are part of the enzyme cocktails that are used for the industrial breakdown of lignocellulosic biomass. Nevertheless, functional studies of GH6 CBHs are scarce and focused on a few model enzymes. To elucidate functional breadth among GH6 CBHs, we conducted a comparative biochemical study of seven GH6 CBHs originating from fungi living in different habitats, in addition to one enzyme variant. The enzyme sequences were investiga...
Source: Biochemical Journal - August 5, 2019 Category: Biochemistry Authors: Christensen, S. J., Krogh, K. B. R. M., Spodsberg, N., Borch, K., Westh, P. Tags: Research Articles Source Type: research

Large-scale chromatin organisation in interphase, mitosis and meiosis
The spatial configuration of chromatin is fundamental to ensure any given cell can fulfil its functional duties, from gene expression to specialised cellular division. Significant technological innovations have facilitated further insights into the structure, function and regulation of three-dimensional chromatin organisation. To date, the vast majority of investigations into chromatin organisation have been conducted in interphase and mitotic cells leaving meiotic chromatin relatively unexplored. In combination, cytological and genome-wide contact frequency analyses in mammalian germ cells have recently demonstrated that ...
Source: Biochemical Journal - August 5, 2019 Category: Biochemistry Authors: MacGregor, I. A., Adams, I. R., Gilbert, N. Tags: Review Articles Source Type: research

Structural insights into SUMO E1-E2 interactions in Arabidopsis uncovers a distinctive platform for securing SUMO conjugation specificity across evolution
SUMOylation of proteins involves the concerted action of the E1-activating enzyme, E2-conjugating enzyme and E3-ligases. An essential discrimination step in the SUMOylation pathway corresponds to the initial interaction between E1 ubiquitin-fold domain (UFD) and E2 enzymes. Although E2 orthologs possess high sequence identity, the E2 binding region of the UFD domains has diverged across evolution. Moreover, in reciprocal in vitro conjugation reactions Arabidopsis E1 and E2 SCE1 fail to interact efficiently with cognate human E2 Ubc9 and E1 partners, respectively. To gain more insights into the properties of this interface ...
Source: Biochemical Journal - July 31, 2019 Category: Biochemistry Authors: Liu, B., Lois, L. M., Reverter, D. Tags: Research Articles Source Type: research

Sugar beet hemoglobins: reactions with nitric oxide and nitrite reveal differential roles for nitrogen metabolism
In contrast with human hemoglobin (Hb) in red blood cells, plant Hbs do not transport oxygen, instead research points towards nitrogen metabolism. Using comprehensive and integrated biophysical methods we characterized three sugar beet Hbs: BvHb1.1, BvHb1.2 and BvHb2. Their affinities for oxygen, CO, and hexacoordination were determined. Their role in nitrogen metabolism was studied by assessing their ability to bind NO, to reduce nitrite (NiR, nitrite reductase), and to form nitrate (NOD, NO dioxygenase). Results show that BvHb1.2 has high NOD-like activity, in agreement with the high nitrate levels found in seeds where t...
Source: Biochemical Journal - July 31, 2019 Category: Biochemistry Authors: Leiva Eriksson, N., Reeder, B. J., Wilson, M. T., Bülow, L. Tags: Research Articles Source Type: research

The role of PTB domain containing adaptor proteins on PICALM-mediated APP endocytosis and localization
One hallmark of Alzheimer's disease (AD) is the presence of amyloid plaques, which mainly consist of the amyloid precursor protein (APP) cleavage product amyloid β (Aβ). For cleavage to occur, the APP must be endocytosed from the cell surface. The phosphatidylinositol binding clathrin assembly protein (PICALM) is involved in clathrin-mediated endocytosis and polymorphisms in and near the gene locus were identified as genetic risk factors for AD. PICALM overexpression enhances APP internalization and Aβ production. Furthermore, PICALM shuttles into the nucleus, but its function within the nucleus is still unk...
Source: Biochemical Journal - July 31, 2019 Category: Biochemistry Authors: Merthan, L., Haller, A., Thal, D. R., von Einem, B., von Arnim, C. A. F. Tags: Research Articles Source Type: research

Structural basis of glycogen metabolism in bacteria
We describe and discuss the remarkable progress made in the understanding of the molecular mechanisms of substrate recognition and product release, allosteric regulation and catalysis of all those enzymes. (Source: Biochemical Journal)
Source: Biochemical Journal - July 31, 2019 Category: Biochemistry Authors: Cifuente, J. O., Comino, N., Trastoy, B., D'Angelo, C., Guerin, M. E. Tags: Review Articles Source Type: research

Biochemical features of primary cells from a pediatric patient with a gain-of-function ODC1 genetic mutation
In conclusion, our patient and potentially other patients that carry a similar ODC1 gain-of-function mutation might benefit from treatment with DFMO, a drug with a good safety profile, to suppress the exceptionally high ODC activity and putrescine levels in the body. (Source: Biochemical Journal)
Source: Biochemical Journal - July 24, 2019 Category: Biochemistry Authors: Schultz, C. R., Bupp, C. P., Rajasekaran, S., Bachmann, A. S. Tags: Research Articles Source Type: research

Comprehensive analysis of yeast ESCRT-III composition in single ESCRT-III deletion mutants
The endosomal sorting complex required for transport (ESCRT)-III is associated with a multitude of cellular processes involving membrane remodeling and abscission. The exact composition of ESCRT-III and the contribution of individual ESCRT-III family members to these diverse functions is unclear. Most of the currently available information about ESCRT-III was obtained with tagged, largely non-functional proteins, which may not correctly reflect the in vivo situation. Here, we performed a comprehensive biochemical analysis of ESCRT-III localization and composition in yeast under purely native conditions. Most of our finding...
Source: Biochemical Journal - July 24, 2019 Category: Biochemistry Authors: Heinzle, C., Mücke, L., Brune, T., Kölling, R. Tags: Research Articles Source Type: research

Less photoprotection can be good in some genetic and environmental contexts
Antioxidant systems modulate oxidant-based signaling networks and excessive removal of oxidants can prevent beneficial acclimation responses. Evidence from mutant, transgenic, and locally adapted natural plant systems is used to interpret differences in the capacity for antioxidation and formulate hypotheses for future inquiry. We focus on the first line of chloroplast antioxidant defense, pre-emptive thermal dissipation of excess absorbed light (monitored as nonphotochemical fluorescence quenching, NPQ) as well as on tocopherol-based antioxidation. Findings from NPQ-deficient and tocopherol-deficient mutants that exhibite...
Source: Biochemical Journal - July 18, 2019 Category: Biochemistry Authors: Demmig-Adams, B., Stewart, J. J., Adams, W. W. Tags: Review Articles Source Type: research

The Mycobacterium tuberculosis capsule: a cell structure with key implications in pathogenesis
Bacterial capsules have evolved to be at the forefront of the cell envelope, making them an essential element of bacterial biology. Efforts to understand the Mycobacterium tuberculosis (Mtb) capsule began more than 60 years ago, but the relatively recent development of mycobacterial genetics combined with improved chemical and immunological tools have revealed a more refined view of capsule molecular composition. A glycogen-like α-glucan is the major constituent of the capsule, with lower amounts of arabinomannan and mannan, proteins and lipids. The major Mtb capsular components mediate interactions with phagocytes t...
Source: Biochemical Journal - July 18, 2019 Category: Biochemistry Authors: Kalscheuer, R., Palacios, A., Anso, I., Cifuente, J., Anguita, J., Jacobs, W. R., Guerin, M. E., Prados-Rosales, R. Tags: Review Articles Source Type: research

Complementary substrate specificity and distinct quaternary assembly of the Escherichia coli aerobic and anaerobic {beta}-oxidation trifunctional enzyme complexes
The trifunctional enzyme (TFE) catalyzes the last three steps of the fatty acid β-oxidation cycle. Two TFEs are present in Escherichia coli, EcTFE and anEcTFE. EcTFE is expressed only under aerobic conditions, whereas anEcTFE is expressed also under anaerobic conditions, with nitrate or fumarate as the ultimate electron acceptor. The anEcTFE subunits have higher sequence identity with the human mitochondrial TFE (HsTFE) than with the soluble EcTFE. Like HsTFE, here it is found that anEcTFE is a membrane-bound complex. Systematic enzyme kinetic studies show that anEcTFE has a preference for medium- and long-chain enoyl...
Source: Biochemical Journal - July 15, 2019 Category: Biochemistry Authors: Sah-Teli, S. K., Hynönen, M. J., Schmitz, W., Geraets, J. A., Seitsonen, J., Pedersen, J. S., Butcher, S. J., Wierenga, R. K., Venkatesan, R. Tags: Research Articles Source Type: research

Structural basis for histone H3K4me3 recognition by the N-terminal domain of the PHD finger protein Spp1
Saccharomyces cerevisiae Spp1, a plant homeodomain (PHD) finger containing protein, is a critical subunit of the histone H3K4 methyltransferase complex of proteins associated with Set1 (COMPASS). The chromatin binding affinity of the PHD finger of Spp1 has been proposed to modulate COMPASS activity. During meiosis, Spp1 plays another role in promoting programmed double-strand break (DSB) formation by binding H3K4me3 via its PHD finger and interacting with a DSB protein, Mer2. However, how the Spp1 PHD finger performs site-specific readout of H3K4me3 is still not fully understood. In the present study, we determined the cry...
Source: Biochemical Journal - July 15, 2019 Category: Biochemistry Authors: He, C., Liu, N., Xie, D., Liu, Y., Xiao, Y., Li, F. Tags: Research Articles Source Type: research

The catalytic mechanism for NO production by the mitochondrial enzyme, sulfite oxidase
Recently, Guenter Schwarz and colleagues published an elegant study in the Biochemical Journal (2019) 476, 1805–1815 which combines kinetic and spectroscopic studies with protein engineering to provide a mechanism for sulfite oxidase (SO)-catalyzed nitrite reduction that yields nitric oxide (NO). This work is noteworthy as it demonstrates that (i) for NO generation, both sulfite and nitrite must bind to the same molybdenum (Mo) center; (ii) upon sulfite reduction, Mo is reduced from +6 (MoVI) to +4 (MoIV) and MoIV reduces nitrite to NO yielding MoV; (iii) the heme moiety, linked to the Mo-center by an 11 amino acid r...
Source: Biochemical Journal - July 15, 2019 Category: Biochemistry Authors: Mutus, B. Tags: Commentaries Source Type: research

Metabolic remodeling of cardiomyocytes identified in phosphoinositide-dependent kinase 1-deficient mice
Metabolic remodeling plays an essential role in the pathophysiology of heart failure (HF). Many studies have shown that the disruption of phosphoinositide-dependent protein kinase-1 (PDK1) caused severe and lethal HF; however, the metabolic pattern of PDK1 deletion remains ambiguous. 1H nuclear magnetic resonance-based metabolomics was applied to explore the altered metabolic pattern in Pdk1-deficient mice. Principle component analysis showed significant separation as early as 4 weeks of age, and dysfunction of metabolism precedes a morphological change in Pdk1-deficient mice. A time trajectory plot indicated that disturbe...
Source: Biochemical Journal - July 9, 2019 Category: Biochemistry Authors: Li, C., Niu, Y., Zheng, H., Shan, C., Chen, Q., Yang, Z., Zhao, L., Yang, C., Gao, H. Tags: Research Articles Source Type: research

Mechanisms of lipid droplet biogenesis
Lipid droplets (LDs) are organelles that compartmentalize nonbilayer-forming lipids in the aqueous cytoplasm of cells. They are ubiquitous in most organisms, including in animals, protists, plants and microorganisms. In eukaryotes, LDs are believed to be derived by a budding and scission process from the surface of the endoplasmic reticulum, and this occurs concomitantly with the accumulation of neutral lipids, most often triacylglycerols and steryl esters. Overall, the mechanisms underlying LD biogenesis are difficult to generalize, in part because of the involvement of different sets of both evolutionarily conserved and ...
Source: Biochemical Journal - July 9, 2019 Category: Biochemistry Authors: Chapman, K. D., Aziz, M., Dyer, J. M., Mullen, R. T. Tags: Review Articles Source Type: research

Correction: Ezrin-anchored PKA phosphorylates serine 369 and 373 on connexin 43 to enhance gap junction assembly, communication, and cell fusion
(Source: Biochemical Journal)
Source: Biochemical Journal - July 2, 2019 Category: Biochemistry Authors: Dukic, A. R., Gerbaud, P., Guibourdenche, J., Thiede, B., Tasken, K., Pidoux, G. Tags: Correction Source Type: research

Nitration-induced ubiquitination and degradation control quality of ERK1
The mitogen-activated protein kinase ERK1/2 (ERKs, extracellular-regulated protein kinases) plays important roles in a wide spectrum of cellular processes and have been implicated in many disease states. The spatiotemporal regulation of ERK activity has been extensively studied. However, scarce information has been available regarding the quality control of the kinases to scavenge malfunctioning ERKs. Using site-specific mutagenesis and mass spectrometry, we found that the disruption of the conserved H-bond between Y210 and E237 of ERK1 through point mutation at or naturally occurring nitration on Y210 initiates a quality ...
Source: Biochemical Journal - July 2, 2019 Category: Biochemistry Authors: Zhang, Y., Huang, X., Wang, J., Wang, X., Liu, X., Chen, Y., Xu, W., Wang, Y. Tags: Research Articles Source Type: research