Sensitive detection of SARS ‐CoV‐2 main protease 3CLpro with an engineered ribonuclease zymogen
AbstractAlongside vaccines and antiviral therapeutics, diagnostic tools are a crucial aid in combating the COVID-19 pandemic caused by the etiological agent SARS-CoV-2. All common assays for infection rely on the detection of viral sub-components, including structural proteins of the virion or fragments of the viral genome. Selective pressure imposed by human intervention of COVID-19 can, however, induce viral mutations that decrease the sensitivity of diagnostic assays based on biomolecular structure, leading to an increase in false-negative results. In comparison, mutations are unlikely to alter thefunction of viral prot...
Source: Protein Science - March 19, 2024 Category: Biochemistry Authors: Evans C. Wralstad, Ronald T. Raines Tags: RESEARCH ARTICLE Source Type: research

Synthesis of site ‐specific Fab‐drug conjugates using ADP‐ribosyl cyclases
AbstractTargeted delivery of small-molecule drugs via covalent attachments to monoclonal antibodies has proved successful in clinic. For this purpose, full-length antibodies are mainly used as drug-carrying vehicles. Despite their flexible conjugation sites and versatile biological activities, intact immunoglobulins with conjugated drugs, which feature relatively large molecular weights, tend to have restricted tissue distribution and penetration and low fractions of payloads. Linking small-molecule therapeutics to other formats of antibody may lead to conjugates with optimal properties. Here, we designed and synthesized A...
Source: Protein Science - March 19, 2024 Category: Biochemistry Authors: Hyo Sun Kim, Kimia Hariri, Xiao ‐Nan Zhang, Liang‐Chieh Chen, Benjamin B. Katz, Hua Pei, Stan G. Louie, Yong Zhang Tags: RESEARCH ARTICLE Source Type: research

HadBD dehydratase from Mycobacterium tuberculosis fatty acid synthase type II: A singular structure for a unique function
AbstractWorldwide, tuberculosis is the second leading infectious killer and multidrug resistance severely hampers disease control. Mycolic acids are a unique category of lipids that are essential for viability, virulence, and persistence of the causative agent,Mycobacterium tuberculosis (Mtb). Therefore, enzymes involved in mycolic acid biosynthesis represent an important class of drug targets. We previously showed that the (3R)-hydroxyacyl-ACP dehydratase (HAD) protein HadD is dedicated mainly to the production of ketomycolic acids and plays a determinant role inMtb biofilm formation and virulence. Here, we discovered tha...
Source: Protein Science - March 19, 2024 Category: Biochemistry Authors: Pascaline Bories, Julie Rima, Samuel Tranier, Julien Marcoux, Yasmina Grimoire, Mathilde Tomaszczyk, Anne Launay, Karine Fata, Hedia Marrakchi, Odile Burlet ‐Schiltz, Lionel Mourey, Manuelle Ducoux‐Petit, Fabienne Bardou, Cécile Bon, An Tags: RESEARCH ARTICLE Source Type: research

Mechanosensitive channel MscL gating transitions coupling with constriction point shift
AbstractThe mechanosensitive channel of large conductance (MscL) acts as an “emergency release valve” that protects bacterial cells from acute hypoosmotic stress, and it serves as a paradigm for studying the mechanism underlying the transduction of mechanical forces. MscL gating is proposed to initiate with an expansion without opening, followed by subsequent pore openi ng via a number of intermediate substates, and ends in a full opening. However, the details of gating process are still largely unknown. Using in vivo viability assay, single channel patch clamp recording, cysteine cross-linking, and tryptophan fluoresc...
Source: Protein Science - March 19, 2024 Category: Biochemistry Authors: Mingfeng Zhang, Siyang Tang, Xiaomin Wang, Sanhua Fang, Yuezhou Li Tags: RESEARCH ARTICLE Source Type: research

Correction to “A novel violet fluorescent protein contains a unique oxidized tyrosine as the simplest chromophore ever reported in fluorescent proteins”
(Source: Protein Science)
Source: Protein Science - March 19, 2024 Category: Biochemistry Tags: CORRECTION Source Type: research

Detection of insulin oligomeric forms by a novel surface plasmon resonance ‐diffusion coefficient based approach
AbstractInsulin is commonly used to treat diabetes and undergoes aggregation at the site of repeated injections in diabetic patients. Moreover, aggregation is also observed during its industrial production and transport and should be avoided to preserve its bioavailability to correctly adjust glucose levels in diabetic patients. However, monitoring the effect of various parameters (pH, protein concentration, metal ions, etc.) on the insulin aggregation and oligomerization state is very challenging. In this work, we have applied a novel Surface Plasmon Resonance (SPR)-based experimental approach to insulin solutions at vari...
Source: Protein Science - March 19, 2024 Category: Biochemistry Authors: Damiano Calcagno, Maria Luisa Perina, Gabriele Antonio Zingale, Irene Pandino, Nunzio Tuccitto, Valentina Oliveri, Maria Cristina Parravano, Giuseppe Grasso Tags: RESEARCH ARTICLE Source Type: research

Structural dynamics and functional cooperativity of human NQO1 by ambient temperature serial crystallography and simulations
AbstractThe human NQO1 (hNQO1) is a flavin adenine nucleotide (FAD)-dependent oxidoreductase that catalyzes the two-electron reduction of quinones to hydroquinones, being essential for the antioxidant defense system, stabilization of tumor suppressors, and activation of quinone-based chemotherapeutics. Moreover, it is overexpressed in several tumors, which makes it an attractive cancer drug target. To decipher new structural insights into the flavin reductive half-reaction of the catalytic mechanism of hNQO1, we have carried serial crystallography experiments at new ID29 beamline of the ESRF to determine, to the best of ou...
Source: Protein Science - March 19, 2024 Category: Biochemistry Authors: Alice Grieco, Sergio Boneta, Jos é A. Gavira, Angel L. Pey, Shibom Basu, Julien Orlans, Daniele de Sanctis, Milagros Medina, Jose Manuel Martin‐Garcia Tags: RESEARCH ARTICLE Source Type: research

Ca2+/CaM dependent protein kinase II (CaMKII) α and CaMKIIβ hub domains adopt distinct oligomeric states and stabilities
AbstractCa2+/calmodulin-dependent protein kinase II (CaMKII) is a multidomain serine/threonine kinase that plays important roles in the brain, heart, muscle tissue, and eggs/sperm. The N-terminal kinase and regulatory domain is connected by a flexible linker to the C-terminal hub domain. The hub domain drives the oligomeric organization of CaMKII, assembling the kinase domains into high local concentration. Previous structural studies have shown multiple stoichiometries of the holoenzyme as well as the hub domain alone. Here, we report a comprehensive study of the hub domain stoichiometry and stability in solution. We solv...
Source: Protein Science - March 19, 2024 Category: Biochemistry Authors: Can Özden, Sara MacManus, Ruth Adafia, Alfred Samkutty, Ana P. Torres‐Ocampo, Scott C. Garman, Margaret M. Stratton Tags: RESEARCH ARTICLE Source Type: research

Reengineering of a flavin ‐binding fluorescent protein using ProteinMPNN
AbstractRecent advances in machine learning techniques have led to development of a number of protein design and engineering approaches. One of them, ProteinMPNN, predicts an amino acid sequence that would fold and match user-defined backbone structure. Its performance was previously tested for proteins composed of standard amino acids, as well as for peptide- and protein-binding proteins. In this short report, we test whether ProteinMPNN can be used to reengineer a non-proteinaceous ligand-binding protein, flavin-based fluorescent protein CagFbFP. We fixed the native backbone conformation and the identity of 20 amino acid...
Source: Protein Science - March 19, 2024 Category: Biochemistry Authors: Andrey Nikolaev, Alexander Kuzmin, Elena Markeeva, Elizaveta Kuznetsova, Yury L. Ryzhykau, Oleg Semenov, Arina Anuchina, Alina Remeeva, Ivan Gushchin Tags: RESEARCH NOTE Source Type: research

Examining evolutionary scale modeling ‐derived different‐dimensional embeddings in the antimicrobial peptide classification through a KNIME workflow
AbstractMolecular features play an important role in different bio-chem-informatics tasks, such as the Quantitative Structure –Activity Relationships (QSAR) modeling. Several pre-trained models have been recently created to be used in downstream tasks, either by fine-tuning a specific model or by extracting features to feed traditional classifiers. In this regard, a new family of Evolutionary Scale Modeling models (terme d as ESM-2 models) was recently introduced, demonstrating outstanding results in protein structure prediction benchmarks. Herein, we studied the usefulness of the different-dimensional embeddings derived...
Source: Protein Science - March 19, 2024 Category: Biochemistry Authors: Karla L. Mart ínez‐Mauricio, César R. García‐Jacas, Greneter Cordoves‐Delgado Tags: TOOLS FOR PROTEIN SCIENCE Source Type: research

Calcium mediated static and dynamic allostery in S100A12: Implications for target recognition by S100 proteins
AbstractStructure and functions of S100 proteins are regulated by two distinct calcium binding EF hand motifs. In this work, we used solution-state NMR spectroscopy to investigate the cooperativity between the two calcium binding sites and map the allosteric changes at the target binding site. To parse the contribution of the individual calcium binding events, variants of S100A12 were designed to selectively bind calcium to either the EF-I (N63A) or EF-II (E31A) loop, respectively. Detailed analysis of the backbone chemical shifts for wildtype protein and its mutants indicates that calcium binding to the canonical EF-II lo...
Source: Protein Science - March 19, 2024 Category: Biochemistry Authors: Qian Wang, Christopher DiForte, Aleksey Aleshintsev, Gianna Elci, Shibani Bhattacharya, Angelo Bongiorno, Rupal Gupta Tags: RESEARCH ARTICLE Source Type: research

Construction of a versatile fusion protein for targeted therapy and immunotherapy
In this study, we expanded the application of antibodies (Abs) by constructing a fusion protein as a versatile tool for Ab-based target cell detection, delivery, and therapy. We first constructed a SpaC Catcher (SpaCC for short) fusion protein that included the C domains of Staphylococcal protein A (SpaC) and the SpyCatcher. SpaCC conjugated with SpyTag-X (S-X) to form the SpaCC-S-X complex, which binds non-covalently to an Ab to form the Ab-SpaCC-S-X protein complex. The “X” can be a variety of small molecules such as fluoresceins, cell-penetrating peptide TAT, Monomethyl auristatin E (MMAE), and DNA. We found that Ab...
Source: Protein Science - March 19, 2024 Category: Biochemistry Authors: Xiu ‐Song Huang, Li‐Ting Yang, Ke Yang, Hang Zhou, Tuersunayi Abudureheman, Wei‐Wei Zheng, Kai‐Ming Chen, Cai‐Wen Duan Tags: RESEARCH ARTICLE Source Type: research

Engineering of IF1 ‐susceptive bacterial F1‐ATPase
AbstractIF1, an inhibitor protein of mitochondrial ATP synthase, suppresses ATP hydrolytic activity of F1. One of the unique features of IF1 is the selective inhibition in mitochondrial F1 (MF1); it inhibits catalysis of MF1 but does not affect F1 with bacterial origin despite high sequence homology between MF1 and bacterial F1. Here, we aimed to engineer thermophilicBacillus F1 (TF1) to confer the susceptibility to IF1 for elucidating the molecular mechanism of selective inhibition of IF1. We first examined the IF1-susceptibility of hybrid F1s, composed of each subunit originating frombovine MF1 (bMF1) or TF1. It was clea...
Source: Protein Science - March 19, 2024 Category: Biochemistry Authors: Yuichiro C. Hatasaki, Ryohei Kobayashi, Ryo R. Watanabe, Mayu Hara, Hiroshi Ueno, Hiroyuki Noji Tags: RESEARCH ARTICLE Source Type: research

Identification of a novel DNA repair inhibitor using an in  silico driven approach shows effective combinatorial activity with genotoxic agents against multidrug‐resistant Escherichia coli
This study offers a novel approach for the potential development of new compounds for use as adjuvants in antineoplastic therapy. (Source: Protein Science)
Source: Protein Science - March 19, 2024 Category: Biochemistry Authors: Lorenzo Bernacchia, Antoine Paris, Arya Gupta, Robert J. Charman, Jake McGreig, Mark N. Wass, Neil M. Kad Tags: RESEARCH ARTICLE Source Type: research

Unraveling the underlying mechanisms of reduced amyloidogenic properties in human calcitonin via double mutations
This study delves into the oligomerization and helical structure formation of the hCT double mutant (Y12LN17H hCT, noted as DM hCT), as well as two single mutants (Y12L and N17H), aiming to elucidate the mechanism behind hCT fibrillization. In addition, computational prediction tools were employed again to identify potential substitutes. Although the results yielded were not entirely satisfactory, a comparison between the newly examined and previously found hCT double mutants provides insights into the reduced aggregation propensity of the latter. This research endeavor holds the promise of informing the design of more eff...
Source: Protein Science - March 19, 2024 Category: Biochemistry Authors: Yu ‐Pei Chang, Pei‐Chun Pan, Ling‐Hsien Tu Tags: RESEARCH ARTICLE Source Type: research