Biochemistry of fluoroprolines: the prospect of making fluorine a bioelement

• Vladimir Kubyshkin,
• Rebecca Davis and
• Nediljko Budisa

Beilstein J. Org. Chem. 2021, 17, 439–460, doi:10.3762/bjoc.17.40

• . We first recapitulate the complex position and biochemical fate of proline in the biochemistry of a cell, discuss the physicochemical properties of fluoroprolines, and overview the attempts to use these amino acids as proline replacements in studies of protein production and folding. Fluorinated

• proline replacements are able to elevate the protein expression speed and yields and improve the thermodynamic and kinetic folding profiles of individual proteins. In this context, fluoroprolines can be viewed as useful tools in the biotechnological toolbox. As a prospect, we envision that proteome-wide

• properties. These factors may translate into an altered structure and stability of a protein containing a fluorinated fragment. What consequences fluorination would have regarding the fitness and survival of the organism relying on fluorine-containing proteins remains an open question. In this context, the

Regioselective chemoenzymatic syntheses of ferulate conjugates as chromogenic substrates for feruloyl esterases

• Olga Gherbovet,
• Fernando Ferreira,
• Apolline Clément,
• Mélanie Ragon,
• Julien Durand,
• Sophie Bozonnet,
• Michael J. O'Donohue and
• Régis Fauré

Beilstein J. Org. Chem. 2021, 17, 325–333, doi:10.3762/bjoc.17.30

• free 4NTC by sodium periodate (Figure 2B). The specific activity (SA) of AnFaeA on 4NTC–linker–Fe (12) was determined to be 3 IU/mg of the protein (with IU corresponding to the international unit of the Fae hydrolytic activity), a value comparable to that measured on destarched wheat bran (3 IU/mg) [44

• µL of 2 M Na2CO3 after 5 min. The OD values at 530 nm were recorded on an Infinite M200 PRO (TECAN) microplate reader. One international unit (IU) of Fae specific activity (SA, expressed in µmol/min/mg or IU/mg) corresponds to the amount of released 4NTC (in µmol) per minute per milligram of protein

¹⁹F NMR as a tool in chemical biology

• Diana Gimenez,
• Aoife Phelan,
• Cormac D. Murphy and
• Steven L. Cobb

Beilstein J. Org. Chem. 2021, 17, 293–318, doi:10.3762/bjoc.17.28

• fluorinated probes and their incorporation into macromolecules, the application of 19F NMR to monitor protein–protein interactions, protein–ligand interactions, physiologically relevant ions and in the structural analysis of proteins and nucleic acids. The continued relevance of the technique to investigate

• biosynthesis and biodegradation of fluorinated organic compounds is also described. Keywords: biotransformation; chemical biology; fluorine; 19F NMR; probes; protein structure; Introduction Although fluorine is abundant in the environment, it is not a nutrient nor is it a feature of biochemistry for most

• another. With regards to probing both the structure and the interactions between biomolecules in complex settings the analytical tool 19F NMR has become invaluable. Some key highlights of how 19F NMR has been employed in this area are given in the following section. Recent advances in protein 19F

The preparation and properties of 1,1-difluorocyclopropane derivatives

• Kymbat S. Adekenova,
• Peter B. Wyatt and
• Sergazy M. Adekenov

Beilstein J. Org. Chem. 2021, 17, 245–272, doi:10.3762/bjoc.17.25

Supramolecular polymerization of sulfated dendritic peptide amphiphiles into multivalent L-selectin binders

• David Straßburger,
• Svenja Herziger,
• Katharina Huth,
• Moritz Urschbach,
• Rainer Haag and
• Pol Besenius

Beilstein J. Org. Chem. 2021, 17, 97–104, doi:10.3762/bjoc.17.10

• [6][7][8][9] are only few of the numerous examples for multivalent protein–protein or protein–carbohydrate interactions that underline their pivotal role in biology. Mimicking polyvalency using synthetic systems has therefore become a growing field and the high degree of functionality renders

• of multivalent interactions is the extracellular adhesion protein L-selectin. L-Selectin plays a critical role in inflammation processes by supporting the migration of leukocytes to inflammatory sites via adhesion to endothelial cells [19][20][21]. On a molecular level, a cationic binding site [22

Molecular basis for protein–protein interactions

• Brandon Charles Seychell and
• Tobias Beck

Beilstein J. Org. Chem. 2021, 17, 1–10, doi:10.3762/bjoc.17.1

• knowledge of protein–protein interactions, common characterisation methods to characterise them, and their role in protein complex formation with some examples. A deep understanding of protein–protein interactions and their molecular interactions is important for a number of applications, including drug

• design. Protein–protein interactions and their discovery are thus an interesting avenue for understanding how protein complexes, which make up the majority of proteins, work. Keywords: characterisation methods; heterooligomeric complex; homooligomeric complex; molecular interactions; protein–protein

• interactions; Introduction From signalling over transport to catalysis, the broad functionality of proteins is essential in the cellular machinery. To this effect, proteins can be seen as the workforce of the cell. Proteins relay some of their functionality via interactions between protein nodes called

Semiautomated glycoproteomics data analysis workflow for maximized glycopeptide identification and reliable quantification

• Steffen Lippold,
• Arnoud H. de Ru,
• Jan Nouta,
• Peter A. van Veelen,
• Magnus Palmblad,
• Manfred Wuhrer and
• Noortje de Haan

Beilstein J. Org. Chem. 2020, 16, 3038–3051, doi:10.3762/bjoc.16.253

• quantification. Keywords: bioinformatics; cysteine oxidation; glycoproteomics; immunoglobulins; mass spectrometry; Introduction Protein glycosylation mainly occurs in the form of N- and O-glycosylation. N-Glycans are attached to Asn within an amino acid consensus sequence (Asn-Xxx-Ser/Thr, Xxx ≠ Pro) and O

• -glycans are attached to Ser or Thr. Glycan compositions can range from monosaccharides (e.g., Tn antigen for O-glycans [1]) to large polysaccharides (e.g., N-glycans of recombinant human erythropoietin [2]). The most common building blocks of human protein glycans are hexoses (glucose, galactose, and

• features, such as the linkage position and anomeric configuration, make protein glycosylation a highly complex posttranslational modification (PTM). Glycoproteomics has become important for many life science disciplines, in particular for biomedical and biopharmaceutical research [3][4][5]. Glycopeptide

Secondary metabolites of Bacillus subtilis impact the assembly of soil-derived semisynthetic bacterial communities

• Heiko T. Kiesewalter,
• Carlos N. Lozano-Andrade,
• Mikael L. Strube and
• Ákos T. Kovács

Beilstein J. Org. Chem. 2020, 16, 2983–2998, doi:10.3762/bjoc.16.248

• , enzyme inhibition, or disruption of bacterial protein synthesis. This knowledge was primarily acquired in vitro when B. subtilis was competing with other microbial monocultures. However, our understanding of the true ecological role of these small molecules is limited. In this study, we have established

• ). For the biosynthesis of B. subtilis NRPs, the phosphopantetheinyl transferase Sfp is needed since it has been shown to activate the peptidyl carrier protein domains, converting it from the inactive apo-form to the active holo-form [37]. B. subtilis has four sfp-dependent SMs, of which three are

• primarily against various filamentous fungi [47][48][49][50][51]. The broad-spectrum antibiotic bacillaene, synthesised by the pksB-S gene cluster, is mainly targeting bacterial protein synthesis [52]. Still, it was also shown that it could protect cells and spores from predation [53]. We recently

Selected peptide-based fluorescent probes for biological applications

• Debabrata Maity

Beilstein J. Org. Chem. 2020, 16, 2971–2982, doi:10.3762/bjoc.16.247

• fluorescence techniques have attracted immense interest. Synthetic peptide-based fluorescent probes are advantageous over protein-based sensors, since they are synthetically accessible, more stable, and can be easily modified in a site-specific manner for selective biological applications. Peptide receptors

• -aminobutyric acid associate with membrane-bound protein receptors and trigger changes in receptor shape and activity with subsequent signaling across the membrane. Noncovalent H-bonding and van der Waals interactions are the basis for the selective molecular recognition between a G-coupled protein receptor and

• membrane-bound receptors interact strongly with short peptidic segments of a larger protein chain, for example, recognition of the RGD sequence (arginine–glycine–aspartic acid; Arg–Gly–Asp) by integrin receptors. They use noncovalent interactions including salt bridges [5]. Vancomycin, a glycopeptide

UV resonance Raman spectroscopy of the supramolecular ligand guanidiniocarbonyl indole (GCI) with 244 nm laser excitation

• Tim Holtum,
• Vikas Kumar,
• Daniel Sebena,
• Jens Voskuhl and
• Sebastian Schlücker

Beilstein J. Org. Chem. 2020, 16, 2911–2919, doi:10.3762/bjoc.16.240

• autofluorescence of the peptide or protein. Here, we demonstrate the use of UVRR spectroscopy with 244 nm laser excitation for the characterization of GCP as well as guanidiniocarbonyl indole (GCI), a next generation supramolecular ligand for the recognition of carboxylates. For demonstrating the feasibility of

• strengths. Various spectroscopic techniques can be employed for monitoring these changes. For example, electronic absorption or fluorescence spectroscopy can probe the spectral differences due to the complexation of the supramolecular ligand with a peptide or protein. However, electronic spectroscopies

• . In earlier studies, we have demonstrated the suitability of UVRR spectroscopy for monitoring the supramolecular binding of monovalent GCP-based ligands with peptides [12][13][14][15][16] and a trivalent GCP-based ligand with the protein leucine zipper, a protein with a single aromatic unit [17], by

Incorporation of a metal-mediated base pair into an ATP aptamer – using silver(I) ions to modulate aptamer function

• Marius H. Heddinga and
• Jens Müller

Beilstein J. Org. Chem. 2020, 16, 2870–2879, doi:10.3762/bjoc.16.236

• aptamer that adopts a guanine quadruplex structure was modified by four pyridine ligands [22]. The addition of Cu(II) or Ni(II) ions leads to the formation of a square-planar complex that reduces the affinity of the modified aptamer to its target protein. To the best of our knowledge, there is no

Encrypting messages with artificial bacterial receptors

• Pragati Kishore Prasad,
• Naama Lahav-Mankovski,
• Leila Motiei and
• David Margulies

Beilstein J. Org. Chem. 2020, 16, 2749–2756, doi:10.3762/bjoc.16.225

• receptors is described. We show that the binding of DNA-based artificial receptors to E. coli expressing His-tagged outer membrane protein C (His-OmpC) induces a Förster resonance energy transfer (FRET) between the dyes, which results in the generation of a unique fluorescence fingerprint. Because the

• reversibly change the properties of the cell. For example, we have shown that synthetic receptors appended with a thiol or a folate group enable bacteria expressing the His-tagged outer membrane protein C (His-OmpC) to bind to gold surfaces or cancer cells, respectively [2]. We have also shown that this

• supplemented with 100 μg/mL of ampicillin at 30 °C. Then, the pre-cultured cells were diluted 1:100 in fresh LB medium supplemented with the same concentration of ampicillin, and incubated until the OD600 reached ≈0.6. In order to induce protein expression, 0.1% rhamnose and 20 μM isopropyl-β-ᴅ-1

Selective recognition of ATP by multivalent nano-assemblies of bisimidazolium amphiphiles through “turn-on” fluorescence response

• Rakesh Biswas,
• Surya Ghosh,
• Shubhra Kanti Bhaumik and
• Supratim Banerjee

Beilstein J. Org. Chem. 2020, 16, 2728–2738, doi:10.3762/bjoc.16.223

• ]. It is a principle ubiquitously used in biology to achieve high affinity binding events with examples ranging from protein–carbohydrate interactions to host–pathogen interactions or cell surface adhesion [12][13][14]. The high affinity originates from the simultaneous interactions of multiple sites in

Enzyme-instructed morphological transition of the supramolecular assemblies of branched peptides

• Dongsik Yang,
• Hongjian He and
• Bing Xu

Beilstein J. Org. Chem. 2020, 16, 2709–2718, doi:10.3762/bjoc.16.221

• noncovalent synthesis [41][42] in a cellular environment. Results and Discussion Molecular design We designed the branched peptides including the DEXXXLLI sequences [43] for this study. The DEXXXLLI (X is any amino acid) sequences are sorting signals of adaptor protein (AP) complexes, which play a critical

• concentrations, exhibit the same morphological appearances, agreeing with the statement that the nanofibers likely are made of Nap-ffky. Cytotoxicity, cell lysates, and protein delivery We investigated the cell compatibility of 1 and 2 by incubation with two kinds of mammalian cells, HeLa and Saos-2 cells, using

• nanofibers in the HeLa cells. Then, we used confocal laser scanning microscopy (CLSM) to examine whether these compounds deliver RPE into HeLa cells. After mixing R-phycoerythrin (RPE) [49], a red fluorescent protein, with 1 or 2 at different concentrations, we incubated the mixtures with HeLa cells for 2 h

Vicinal difluorination as a C=C surrogate: an analog of piperine with enhanced solubility, photostability, and acetylcholinesterase inhibitory activity

• Yuvixza Lizarme-Salas,
• Alexandra Daryl Ariawan,
• Ranjala Ratnayake,
• Hendrik Luesch,
• Angela Finch and
• Luke Hunter

Beilstein J. Org. Chem. 2020, 16, 2663–2670, doi:10.3762/bjoc.16.216

• microenvironment of a protein binding site could also change the relative energies of the various F–C–C–F and F–C–C=O rotamers, offering the possibility that analog 2 might be an effective conformational mimic of 1 in some environments but not in others. Herein, we describe the optimisation of a synthetic route to

Anion exchange resins in phosphate form as versatile carriers for the reactions catalyzed by nucleoside phosphorylases

• Julia N. Artsemyeva,
• Ekaterina A. Remeeva,
• Tatiana N. Buravskaya,
• Irina D. Konstantinova,
• Roman S. Esipov,
• Anatoly I. Miroshnikov,
• Natalia M. Litvinko and
• Igor A. Mikhailopulo

Beilstein J. Org. Chem. 2020, 16, 2607–2622, doi:10.3762/bjoc.16.212

• activity 167 IU per mg protein), uridine phosphorylase (UP, specific activity 140 IU per mg protein in solution 17 mg/mL), and purine nucleoside phosphorylase (PNP, specific activity 27 IU per mg of protein) [80] have been used throughout of the studies. The powdered enzymes were dissolved in 5 mM K

Comparative ligand structural analytics illustrated on variably glycosylated MUC1 antigen–antibody binding

• Christopher B. Barnett,
• Tharindu Senapathi and
• Kevin J. Naidoo

Beilstein J. Org. Chem. 2020, 16, 2540–2550, doi:10.3762/bjoc.16.206

• conformation of the peptide portion of the antigen and does not bind directly. Previous studies have shown that O-glycosylation may provide increased physical stability [20], rigid conformations for protein stability [21], induce the formation of stiff and extended peptide conformations [22], and may affect

• mucin octapeptide showed that the peptide conformation depended on the extent of glycosylation. Glycosylation induces small changes in protein structure and shifts it from a random to a more turn-like structure [26]. Kirnasky et al. noted that O-glycosylation slightly affected the conformational

• server [30] which includes several helper tools (PDB Manipulator [31] and Glycan Reader [32][33]), was used to build these systems and generate input files [34] for use with OpenMM. Five systems were built in CHARMM-GUI based on initial structures from the Protein Data bank (PDB ID:5T6P, 5T78). The

Leveraging glycomics data in glycoprotein 3D structure validation with Privateer

• Haroldas Bagdonas,
• Daniel Ungar and
• Jon Agirre

Beilstein J. Org. Chem. 2020, 16, 2523–2533, doi:10.3762/bjoc.16.204

• convey significantly more information than what is available through protein synthesis and the expression of the genetic code alone. For example, glycosylation is used as a switch to modulate protein activity [1]; glycosylation plays a crucial part in folding/unfolding pathways of some proteins in cells

• [2][3]; the level of N-glycan expression regulates the adhesiveness of a cell [4]; glycosylation also plays a role in immune function [5] and cellular signalling [5][6]. At the forefront, glycosylation plays a significant role in influencing protein–protein interactions. For example, the influenza

• and protein interactions [12]. Unfortunately, significant challenges have affected the determination of glycoprotein structures for decades and have had a detrimental impact on the quality and reliability of the produced models. Anomalies have been reported regarding carbohydrate nomenclature [13

NMR Spectroscopy of supramolecular chemistry on protein surfaces

• Peter Bayer,
• Anja Matena and
• Christine Beuck

Beilstein J. Org. Chem. 2020, 16, 2505–2522, doi:10.3762/bjoc.16.203

• interaction of proteins with their interaction partners, both biomolecules and synthetic ligands. In recent years, the focus in chemistry has kept expanding from targeting small binding pockets in proteins to recognizing patches on protein surfaces, mostly via supramolecular chemistry, with the goal to

• modulate protein–protein interactions. Here we present NMR methods that have been applied to characterize these molecular interactions and discuss the challenges of this endeavor. Keywords: molecular recognition; NMR; protein ligand interaction; protein surfaces; supramolecular chemistry; Introduction In

• recent years, the focus of biochemical research and drug development has shifted from the inhibition of single enzymes to targeting protein-protein interactions [1][2], which play key roles in cellular function and dysfunction [3][4]. Enzymes usually bind their substrates in deep pockets with specific

Computational tools for drawing, building and displaying carbohydrates: a visual guide

• Kanhaya Lal,
• Rafael Bermeo and
• Serge Perez

Beilstein J. Org. Chem. 2020, 16, 2448–2468, doi:10.3762/bjoc.16.199

• formats [11]. Along the same line, the development of various other applications allowed the users to sketch 2D-glycan structures by dragging and dropping monosaccharides to canvas to generate 3D structures for further usages. These depictions comply with protein data bank (PDB) [12] format, or in the

• form of images [13][14]. Besides, these tools for representing glycans in 2D and 3D shape [15] allowed the integration of glycans into protein structures or complexes. The tools developed in the last few years have automated the sketching of glycans and glycopeptides, allowing rapid display of

• hand, the Glycan Modeler allows in silico N-/O-glycosylation for glycan-protein complexes and generates a “most relevant” glycan structure through Glycan Fragment Database (GFDB) [68] search which gives proper orientations relative to the target protein. In the absence of target glycan sequence in GFDB

The B & B approach: Ball-milling conjugation of dextran with phenylboronic acid (PBA)-functionalized BODIPY

• Patrizia Andreozzi,
• Lorenza Tamberi,
• Elisamaria Tasca,
• Gina Elena Giacomazzo,
• Marta Martinez,
• Mirko Severi,
• Marco Marradi,
• Stefano Cicchi,
• Sergio Moya,
• Giacomo Biagiotti and
• Barbara Richichi

Beilstein J. Org. Chem. 2020, 16, 2272–2281, doi:10.3762/bjoc.16.188

• range of biomolecules: amino acids [8], peptides [9], glycosides [10], nucleosides/nucleotides [11], and lipids [12]. Also, protein-based nano-bio-conjugates [13] have been prepared by ball milling, retaining the native properties of the proteins after mechanochemical synthesis. Boronic acids and their

Tools for generating and analyzing glycan microarray data

• Akul Y. Mehta,
• Jamie Heimburg-Molinaro and
• Richard D. Cummings

Beilstein J. Org. Chem. 2020, 16, 2260–2271, doi:10.3762/bjoc.16.187

• the mammalian body. They play a vital role in a number of physiologic and pathologic conditions. Glycan microarrays allow a plethora of information to be obtained on protein–glycan binding interactions. In this review, we describe the intricacies of the generation of glycan microarray data and the

• the dataset. D) Structural information tools 1. GlyMDB: Status: Available. Address: http://www.glycanstructure.org/glymdb/. Description: GlyMDB is a web-based database which links glycan microarray binding data from the CFG database to protein structures (PDB) [51]. A user can select a dataset from

• the CFG dataset available and set thresholds for binding versus nonbinding. The application can then show you motifs which make a significant binding contribution on the microarray. In addition it allows you to quickly search for PDB files with sequence identity matching to the protein sample put on

GlypNirO: An automated workflow for quantitative N- and O-linked glycoproteomic data analysis

• Toan K. Phung,
• Cassandra L. Pegg and
• Benjamin L. Schulz

Beilstein J. Org. Chem. 2020, 16, 2127–2135, doi:10.3762/bjoc.16.180

• Queensland, St. Lucia, QLD 4072, Australia 10.3762/bjoc.16.180 Abstract Mass spectrometry glycoproteomics is rapidly maturing, allowing unprecedented insights into the diversity and functions of protein glycosylation. However, quantitative glycoproteomics remains challenging. We developed GlypNirO, an

• -glycosylation; O-glycosylation; Python; Introduction Glycosylation is a key post-translational modification critical for protein folding and function in eukaryotes [1][2][3]. Diverse types of glycosylation are known, all involving modification of specific amino acid residues with complex carbohydrate

• ]. This heterogeneity can take the form of variable occupancy, also known as macroheterogeneity – the presence or absence of modification at a particular site in a protein, due to inefficient transfer of the initial glycan structure [5]. In addition, the non-template-driven synthesis of glycan structures

Muyocopronones A and B: azaphilones from the endophytic fungus Muyocopron laterale

• Ken-ichi Nakashima,
• Junko Tomida,
• Tomoe Tsuboi,
• Yoshiaki Kawamura and
• Makoto Inoue

Beilstein J. Org. Chem. 2020, 16, 2100–2107, doi:10.3762/bjoc.16.177

• fungus; modified Mosher’s method; Muyocopron laterale; polyketides; Introduction Azaphilones, which are a class of fungal polyketides with diverse structures, have received growing attention due to their various biological activities, such as the inhibition of some protein–protein interactions [1][2

• ], tau aggression [3], and heat shock protein 90 [4], in addition to their antimicrobial, cytotoxic, anticancer, and anti-inflammatory effects [5]. To date, over 400 azaphilones have been reported from various fungal strains, the majority of which have been produced by ascomycetes belonging to the

Clustering and curation of electropherograms: an efficient method for analyzing large cohorts of capillary electrophoresis glycomic profiles for bioprocessing operations

• Ian Walsh,
• Matthew S. F. Choo,
• Sim Lyn Chiin,
• Amelia Mak,
• Shi Jie Tay,
• Pauline M. Rudd,
• Yang Yuansheng,
• Andre Choo,
• Ho Ying Swan and
• Terry Nguyen-Khuong

Beilstein J. Org. Chem. 2020, 16, 2087–2099, doi:10.3762/bjoc.16.176

• purchased from Merck Millipore, (USA). Protein A HP Spintrap (28-9031-32) was purchased from GE Healthcare, USA. FAST Glycan Kit (Part no: B94499PTO, SCIEX, USA). Ammonium formate (Part No. 186007081), RapiGest SF (Part No. 186001860), RapiFluor-MS Reagent Solution (Part No. 186008091), and ACQUITY UPLC

• . Sample preparation of antibody N-glycans Protein A purification and buffer exchange: The collected cell supernatant was filtered through a 0.22 µM PVDF syringe filter (Sterile Millex Filter, Merck Millipore, Ireland). Antibodies were then purified using protein A spin trap columns (Protein A HP Spintrap

• using UPLC-MS The supernatant was purified using Protein A HP SpinTrap (GE Healthcare). The purified glycoprotein obtained was buffer exchanged into water using a 10 kDa molecular weight cut-off filter (Merck Millipore) to eliminate any salts and nucleophiles that could interfere with the subsequent