We present a novel system that leverages curators in the loop to develop a dataset and model for detecting structure features and functional annotations at residue-level from standard publication text. Our approach involves the integration of data from multiple resources, including PDBe, EuropePMC, PubMedCentral, and PubMed, combined with annotation guidelines from UniProt, and LitSuggest and HuggingFace models as tools in the annotation process. A team of seven annotators manually curated ten articles for named entities, which we utilized to train a starting PubmedBert model from HuggingFace. Using a human-in-the-loop annotation system, we iteratively developed the best model with commendable performance metrics of 0.90 for precision, 0.92 for recall, and 0.91 for F1-measure. Our proposed system showcases a successful synergy of machine learning techniques and human expertise in curating a dataset for residue-level functional annotations and protein structure features. The results demonstrate the potential for broader applications in protein research, bridging the gap between advanced machine learning models and the indispensable insights of domain experts.

• MeSH
• databáze proteinů MeSH
• lidé MeSH
• proteiny * chemie   MeSH
• strojové učení * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• dataset MeSH
• Názvy látek
• proteiny * MeSH

The Acyl-CoA-binding domain-containing protein (ACBD3) plays multiple roles across the cell. Although generally associated with the Golgi apparatus, it operates also in mitochondria. In steroidogenic cells, ACBD3 is an important part of a multiprotein complex transporting cholesterol into mitochondria. Balance in mitochondrial cholesterol is essential for proper mitochondrial protein biosynthesis, among others. We generated ACBD3 knock-out (ACBD3-KO) HEK293 and HeLa cells and characterized the impact of protein absence on mitochondria, Golgi, and lipid profile. In ACBD3-KO cells, cholesterol level and mitochondrial structure and functions are not altered, demonstrating that an alternative pathway of cholesterol transport into mitochondria exists. However, ACBD3-KO cells exhibit enlarged Golgi area with absence of stacks and ribbon-like formation, confirming the importance of ACBD3 in Golgi stacking. The glycosylation of the LAMP2 glycoprotein was not affected by the altered Golgi structure. Moreover, decreased sphingomyelins together with normal ceramides and sphingomyelin synthase activity reveal the importance of ACBD3 in ceramide transport from ER to Golgi.

• Klíčová slova
• ACBD3, Golgi, OXPHOS, cholesterol, knock-out, mitochondria,
• MeSH
• adaptorové proteiny signální transdukční metabolismus   MeSH
• biologický transport fyziologie   MeSH
• ceramidy metabolismus   MeSH
• cholesterol metabolismus   MeSH
• glykosylace MeSH
• Golgiho aparát metabolismus   MeSH
• HEK293 buňky MeSH
• HeLa buňky MeSH
• lidé MeSH
• membránové proteiny metabolismus   MeSH
• membránový protein 2 asociovaný s lyzozomy metabolismus   MeSH
• mitochondrie metabolismus   MeSH
• signální transdukce fyziologie   MeSH
• transferasy pro jiné substituované fosfátové skupiny metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• ACBD3 protein, human MeSH Prohlížeč
• adaptorové proteiny signální transdukční MeSH
• ceramidy MeSH
• cholesterol MeSH
• membránové proteiny MeSH
• membránový protein 2 asociovaný s lyzozomy MeSH
• phosphatidylcholine-ceramide phosphocholine transferase MeSH Prohlížeč
• transferasy pro jiné substituované fosfátové skupiny MeSH

Many picornaviruses hijack the Golgi resident Acyl-coenzyme A binding domain containing 3 (ACBD3) protein in order to recruit the phosphatidylinositol 4-kinase B (PI4KB) to viral replication organelles (ROs). PI4KB, once recruited and activated by ACBD3 protein, produces the lipid phosphatidylinositol 4-phosphate (PI4P), which is a key step in the biogenesis of viral ROs. To do so, picornaviruses use their small nonstructural protein 3A that binds the Golgi dynamics domain of the ACBD3 protein. Here, we present the analysis of the highly flexible ACBD3 proteins and the viral 3A protein in solution using small-angle X-ray scattering and computer simulations. Our analysis revealed that both the ACBD3 protein and the 3A:ACBD3 protein complex have an extended and flexible conformation in solution.

• Klíčová slova
• ACBD3, RNA virus, coarse-grained simulations, host factor, intrinsically disordered regions, picornavirus, small-angle X-ray scattering (SAXS),
• MeSH
• acylkoenzym A chemie   metabolismus   MeSH
• adaptorové proteiny signální transdukční chemie   metabolismus   MeSH
• lidé MeSH
• membránové proteiny chemie   metabolismus   MeSH
• Picornaviridae chemie   metabolismus   MeSH
• vazebná místa MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ACBD3 protein, human MeSH Prohlížeč
• acylkoenzym A MeSH
• adaptorové proteiny signální transdukční MeSH
• membránové proteiny MeSH

Enteroviruses, members of the family of picornaviruses, are the most common viral infectious agents in humans causing a broad spectrum of diseases ranging from mild respiratory illnesses to life-threatening infections. To efficiently replicate within the host cell, enteroviruses hijack several host factors, such as ACBD3. ACBD3 facilitates replication of various enterovirus species, however, structural determinants of ACBD3 recruitment to the viral replication sites are poorly understood. Here, we present a structural characterization of the interaction between ACBD3 and the non-structural 3A proteins of four representative enteroviruses (poliovirus, enterovirus A71, enterovirus D68, and rhinovirus B14). In addition, we describe the details of the 3A-3A interaction causing the assembly of the ACBD3-3A heterotetramers and the interaction between the ACBD3-3A complex and the lipid bilayer. Using structure-guided identification of the point mutations disrupting these interactions, we demonstrate their roles in the intracellular localization of these proteins, recruitment of downstream effectors of ACBD3, and facilitation of enterovirus replication. These structures uncovered a striking convergence in the mechanisms of how enteroviruses and kobuviruses, members of a distinct group of picornaviruses that also rely on ACBD3, recruit ACBD3 and its downstream effectors to the sites of viral replication.

• MeSH
• adaptorové proteiny signální transdukční chemie   genetika   metabolismus   MeSH
• fosfotransferasy s alkoholovou skupinou jako akceptorem genetika   metabolismus   MeSH
• HEK293 buňky MeSH
• interakce hostitele a patogenu * MeSH
• konformace proteinů MeSH
• krystalizace MeSH
• krystalografie rentgenová MeSH
• lidé MeSH
• membránové proteiny chemie   genetika   metabolismus   MeSH
• molekulární modely MeSH
• mutace MeSH
• Picornaviridae fyziologie   MeSH
• replikace viru * MeSH
• sekvence aminokyselin MeSH
• sekvenční homologie MeSH
• vazba proteinů MeSH
• virové proteiny chemie   genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ACBD3 protein, human MeSH Prohlížeč
• adaptorové proteiny signální transdukční MeSH
• fosfotransferasy s alkoholovou skupinou jako akceptorem MeSH
• membránové proteiny MeSH
• virové proteiny MeSH

The minor phospholipid, phosphatidylinositol 4-phosphate (PI4P), is emerging as a key regulator of lipid transfer in ER-membrane contact sites. Four different phosphatidylinositol 4-kinase (PI4K) enzymes generate PI4P in different membrane compartments supporting distinct cellular processes, many of which are crucial for the maintenance of cellular integrity but also hijacked by intracellular pathogens. While type III PI4Ks have been targeted by small molecular inhibitors, thus helping decipher their importance in cellular physiology, no inhibitors are available for the type II PI4Ks, which hinders investigations into their cellular functions. Here, we describe the identification of small molecular inhibitors of PI4K type II alpha (PI4K2A) by implementing a large scale small molecule high-throughput screening. A novel assay was developed that allows testing of selected inhibitors against PI4K2A in intact cells using a bioluminescence resonance energy transfer approach adapted to plate readers. The compounds disclosed here will pave the way to the optimization of PI4K2A inhibitors that can be used in cellular and animal studies to better understand the role of this enzyme in both normal and pathological states.

• Klíčová slova
• endosome, phosphoinositide, vesicular traffic,
• MeSH
• 1-fosfatidylinositol-4-kinasa antagonisté a inhibitory   chemie   metabolismus   MeSH
• biologický transport MeSH
• Cercopithecus aethiops MeSH
• COS buňky MeSH
• endozomy účinky léků   metabolismus   MeSH
• Golgiho aparát účinky léků   metabolismus   MeSH
• HEK293 buňky MeSH
• inhibitory enzymů metabolismus   farmakologie   MeSH
• konformace proteinů MeSH
• lidé MeSH
• preklinické hodnocení léčiv MeSH
• rychlé screeningové testy * MeSH
• simulace molekulového dockingu MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Intramural MeSH
• Research Support, U.S. Gov't, Non-P.H.S. MeSH
• Názvy látek
• 1-fosfatidylinositol-4-kinasa MeSH
• inhibitory enzymů MeSH

Phosphatidylinositol 4-kinase IIIβ (PI4KB) is a key enzyme of the Golgi system because it produces its lipid hallmark - the phosphatidylinositol 4-phosphate (PI4P). It is recruited to Golgi by the Golgi resident ACBD3 protein, regulated by 14-3-3 proteins and it also serves as an adaptor because it recruits the small GTPase Rab11. Here, we analyzed the protein complexes formed by PI4KB in vitro using small angle x-ray scattering (SAXS) and we discovered that these protein complexes are highly flexible. The 14-3-3:PI4KB:Rab11 protein complex has 2:1:1 stoichiometry and its different conformations are rather compact, however, the ACBD3:PI4KB protein complex has both, very compact and very extended conformations. Furthermore, in vitro reconstitution revealed that the membrane is necessary for the formation of ACBD3:PI4KB:Rab11 protein complex at physiological (nanomolar) concentrations.

• MeSH
• adaptorové proteiny signální transdukční metabolismus   MeSH
• fosfotransferasy s alkoholovou skupinou jako akceptorem metabolismus   MeSH
• intracelulární membrány metabolismus   MeSH
• maloúhlový rozptyl MeSH
• membránové proteiny metabolismus   MeSH
• multimerizace proteinu * MeSH
• proteiny 14-3-3 metabolismus   MeSH
• Rab proteiny vázající GTP metabolismus   MeSH
• rekombinantní proteiny metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ACBD3 protein, human MeSH Prohlížeč
• adaptorové proteiny signální transdukční MeSH
• fosfotransferasy s alkoholovou skupinou jako akceptorem MeSH
• membránové proteiny MeSH
• phosphatidylinositol 4-kinase IIIbeta, human MeSH Prohlížeč
• proteiny 14-3-3 MeSH
• Rab proteiny vázající GTP MeSH
• rab11 protein MeSH Prohlížeč
• rekombinantní proteiny MeSH

Zika virus is a global health threat due to significantly elevated risk of fetus malformations in infected pregnant women. Currently, neither an effective therapy nor a prophylactic vaccination is available for clinical use, desperately necessitating novel therapeutics and approaches to obtain them. Here, we present a structural model of the Zika virus RNA-dependent RNA polymerase (ZIKV RdRp) in complex with template and nascent RNAs, Mg2+ ions and accessing nucleoside triphosphate. The model allowed for docking studies aimed at effective pre-screening of potential inhibitors of ZIKV RdRp. Applicability of the structural model for docking studies was illustrated with the NITD008 artificial nucleotide that is known to effectively inhibit the function of the ZIKV RdRp. The ZIKV RdRp - RNA structural model is provided for all possible variations of the nascent RNA bases pairs to enhance its general utility in docking and modelling experiments. The developed model makes the rational design of novel nucleosides and nucleotide analogues feasible and thus provides a solid platform for the development of advanced antiviral therapy.

• MeSH
• adenosin analogy a deriváty   chemie   farmakologie   MeSH
• hořčík chemie   MeSH
• infekce virem zika genetika   virologie   MeSH
• konformace proteinů účinky léků   MeSH
• lidé MeSH
• molekulární modely MeSH
• nukleosidy chemie   MeSH
• nukleotidy chemie   MeSH
• polyfosfáty chemie   MeSH
• replikace viru genetika   MeSH
• RNA-dependentní RNA-polymerasa chemie   genetika   MeSH
• RNA chemie   genetika   MeSH
• simulace molekulového dockingu MeSH
• virové nestrukturální proteiny chemie   genetika   MeSH
• virus zika chemie   genetika   patogenita   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• adenosin MeSH
• hořčík MeSH
• NITD008 MeSH Prohlížeč
• nukleosidy MeSH
• nukleotidy MeSH
• polyfosfáty MeSH
• RNA-dependentní RNA-polymerasa MeSH
• RNA MeSH
• triphosphoric acid MeSH Prohlížeč
• virové nestrukturální proteiny MeSH

Phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) is a critically important regulatory lipid of the plasma membrane (PM); however, little is known about how cells regulate PM PI(4,5)P2 levels. Here, we show that the phosphatidylinositol 4-phosphate (PI4P)/phosphatidylserine (PS) transfer activity of the endoplasmic reticulum (ER)-resident ORP5 and ORP8 is regulated by both PM PI4P and PI(4,5)P2 Dynamic control of ORP5/8 recruitment to the PM occurs through interactions with the N-terminal Pleckstrin homology domains and adjacent basic residues of ORP5/8 with both PI4P and PI(4,5)P2 Although ORP5 activity requires normal levels of these inositides, ORP8 is called on only when PI(4,5)P2 levels are increased. Regulation of the ORP5/8 attachment to the PM by both phosphoinositides provides a powerful means to determine the relative flux of PI4P toward the ER for PS transport and Sac1-mediated dephosphorylation and PIP 5-kinase-mediated conversion to PI(4,5)P2 Using this rheostat, cells can maintain PI(4,5)P2 levels by adjusting the availability of PI4P in the PM.

• MeSH
• biologický transport MeSH
• buněčná membrána metabolismus   MeSH
• endoplazmatické retikulum metabolismus   MeSH
• fosfatidylinositol-4,5-difosfát metabolismus   MeSH
• fosfatidylinositolfosfáty metabolismus   MeSH
• fosfatidylseriny metabolismus   MeSH
• fosfotransferasy s alkoholovou skupinou jako akceptorem metabolismus   MeSH
• HEK293 buňky MeSH
• krysa rodu Rattus MeSH
• lidé MeSH
• proteinové domény MeSH
• steroidní receptory chemie   metabolismus   MeSH
• substrátová specifita MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Research Support, N.I.H., Extramural MeSH
• Research Support, N.I.H., Intramural MeSH
• Názvy látek
• fosfatidylinositol-4,5-difosfát MeSH
• fosfatidylinositolfosfáty MeSH
• fosfatidylseriny MeSH
• fosfotransferasy s alkoholovou skupinou jako akceptorem MeSH
• Oxysterol Binding Proteins MeSH
• phosphatidylinositol 4-phosphate MeSH Prohlížeč
• steroidní receptory MeSH

Most single stranded plus RNA viruses hijack phosphatidylinositol 4-kinases (PI4Ks) to generate membranes highly enriched in phosphatidylinositol 4-phosphate (PI4P). These membranous compartments known as webs, replication factories or replication organelles are essential for viral replication because they provide protection from the innate intracellular immune response while serving as platforms for viral replication. Using purified recombinant proteins and biomimetic model membranes we show that the nonstructural viral 3A protein is sufficient to promote membrane hyper-phosphorylation given the proper intracellular cofactors (PI4KB and ACBD3). However, our bio-mimetic in vitro reconstitution assay revealed that rather than the presence of PI4P specifically, negative charge alone is sufficient for the recruitment of 3Dpol enzymes to the surface of the lipid bilayer. Additionally, we show that membrane tethered viral 3B protein (also known as Vpg) works in combination with the negative charge to increase the efficiency of membrane recruitment of 3Dpol.

• MeSH
• adaptorové proteiny signální transdukční genetika   metabolismus   MeSH
• buněčná membrána metabolismus   MeSH
• fosfatidylinositolfosfáty metabolismus   MeSH
• fosfotransferasy s alkoholovou skupinou jako akceptorem genetika   metabolismus   MeSH
• Kobuvirus enzymologie   MeSH
• lidé MeSH
• membránové proteiny genetika   metabolismus   MeSH
• pikornavirové infekce metabolismus   virologie   MeSH
• virové nestrukturální proteiny genetika   metabolismus   MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ACBD3 protein, human MeSH Prohlížeč
• adaptorové proteiny signální transdukční MeSH
• fosfatidylinositolfosfáty MeSH
• fosfotransferasy s alkoholovou skupinou jako akceptorem MeSH
• membránové proteiny MeSH
• phosphatidylinositol 4-kinase IIIbeta, human MeSH Prohlížeč
• phosphatidylinositol 4-phosphate MeSH Prohlížeč
• virové nestrukturální proteiny MeSH

We report on an extensive structure-activity relationship study of novel PI4K IIIβ inhibitors. The purine derivative of the potent screening hit T-00127-HEV1 has served as a suitable starting point for a thorough investigation of positions 8 and 2. While position 8 of the purine scaffold can only bear a small substituent to maintain the inhibitory activity, position 2 is opened for extensive modification and can accommodate even substituted phenyl rings without the loss of PI4K IIIβ inhibitory activity. These empirical observations nicely correlate with the results of our docking study, which suggests that position 2 directs towards solution and can provide the necessary space for the interaction with remote residues of the enzyme, whereas the cavity around position 8 is strictly limited. The obtained compounds have also been subjected to antiviral screening against a panel of (+)ssRNA viruses.

• Klíčová slova
• Antiviral agent, Hepatitis C virus, PI4K IIIβ, Phosphatidylinositol 4-kinase, Purine,
• MeSH
• antivirové látky chemická syntéza   chemie   farmakologie   MeSH
• enterovirus B lidský účinky léků   MeSH
• fosfotransferasy s alkoholovou skupinou jako akceptorem antagonisté a inhibitory   metabolismus   MeSH
• HeLa buňky MeSH
• Hepacivirus účinky léků   MeSH
• inhibitory proteinkinas chemická syntéza   chemie   farmakologie   MeSH
• lidé MeSH
• mikrobiální testy citlivosti MeSH
• molekulární modely MeSH
• molekulární struktura MeSH
• puriny chemická syntéza   chemie   farmakologie   MeSH
• Rhinovirus účinky léků   MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• vztahy mezi strukturou a aktivitou MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antivirové látky MeSH
• fosfotransferasy s alkoholovou skupinou jako akceptorem MeSH
• inhibitory proteinkinas MeSH
• phosphatidylinositol 4-kinase IIIbeta, human MeSH Prohlížeč
• purine MeSH Prohlížeč
• puriny MeSH