Incorporation of Indoylated Phenylalanine Yields a Sub-Micromolar Selective Melanocortin-4 Receptor Antagonist Tetrapeptide. 

M. Ericson, C. Larson, K. Freemann, L. Nicke, A. Geyer, C. Haskell-Luevano. ACS Omega, 2022, in print

Manuscript ID: ao-2022-03307d.R1 

Abstract 

The silica mineralisation properties of synthetic Silaffin-1A1 (synSil‑1A1). 

Fabian Daus, Xiulan Xie, Armin Geyer. Org. Biomol. Chem., 2022, 20, 3387-3396. 

DOI: 10.1039/D2OB00390B

Supplementary Information

Abstract The synthetic monodisperse pentadecapeptide synSil-1A1 is a representative of the microdisperse mixture of the native silaffin natSil-1A1 produced by the diatom Cylindrotheca fusiformis. The octaphosphorylated zwitterionic synSil-1A1 is able to mineralise silica under slightly acidic conditions at pH 5.5, which is the physiologically relevant pH range assumed. Like the posttranslational modifications of the native silaffins, synSil-1A1 is functionalised on all four lysine and phosphorylated on all seven serine residues. We describe the synthesis of a trimethyl-δ-hydroxy-L-lysine building block, the incorporation of this choline-type amino acid in peptide synthesis and its phosphorylation, together with all further posttranslational modifications observed in the native silaffins. Quantitative structure–activity relationships from silicification experiments at high dilution reveal the unique mineralisation properties of the hyperphosphorylated peptide as a single substance and in interaction with long-chain polyamines (LCPA). Diffusion-ordered spectroscopy (DOSY) experiments reveal the formation of polyelectrolyte complexes (PEC) between synSil-1A1 and long-chain polyamines, which promotes the silicification process. The microdroplets have an overall balanced ratio of 100–150 cationic and the same number of anionic charges. The unique zwitterionic synSil-1A1 confirms the prevailing molecular model of biosilicification and validates it with quantitative data based on a single phosphopeptide species, avoiding the usual unphysiologically high concentrations of phosphate of many previous in vitro silicification experiments.

 (no graphical abstract available)

Crystal structure and supramolecularity of (E)-4-(4-tert-butylphenyl)but-3-en-2-one. 

Eman Al-Masri, Mahmoud Al-Refai, Harbi Tomah Al-Masri, Armin Geyer, Sergei I. Ivlev, Basem F. Ali. Jordan Journal of Chemistry 2021, Vol. 16, No. 3, 99-104.

DOI: 10.47014/16.3.1

Synthesis, characterization, antibacterial and cytotoxic evaluation of new 6-(chlorothiophenyl)-2-(2-oxopropoxy)pyridine-3-carbonitrile derivatives and their corresponding furo[2,3-b]pyridines derivatives.

Ala’a B. Said, Mahmoud Al-Refai, Armin Geyer, Iman A. Mansi, Basem Fares Ali. Heterocycles 2021, Vol. 102, No. 11, 2153-2167.

DOI: 10.3987/com-21-14534

Abstract The synthesis of a new series of 6-(chlorothiophenyl)-2-(2-oxopropoxy)pyridine-3-carbonitrile compounds 2 and 5 and their furo[2,3-b]pyridines bearing heteroaryl substituents 3 and 6 in high yield is reported. The 6-(chlorothiophenyl)-2-(2-oxopropoxy)pyridine-3-carbonitrile derivatives (2a-f) and (5a-d) were prepared from the corresponding 3-cyano-(2H)-pyridones (1a-f) and (4a-d) followed by the Thorpe-Ziegler ring cyclization in the presence of sodium methoxide to give the target furo[2,3-b]pyridine derivatives (3a-f) and (6a-d). Proposed structures of the new compounds are based on NMR and mass spectral data. Antibacterial evaluation of (2a-d), (2f), (5a-d), (3a-d), (3f), and (6a-d) derivatives against eight different microorganisms shows no activity towards any of the tested bacteria while compounds (6a) and (6c) showed weak antibacterial activity with inhibition zone of 10 mm. Cytotoxic assessment against MCF7 breast cancer cells reveals that 10 of the derivatives were found to decrease cell viability in a dose dependent manner at concentration of 100 μM, but six of them decreased it to less than 50% and had IC50’s ranging from 23.3 to 41.2 μM.

High five! Methyl probes at five ring positions of phenylalanine explore the hydrophobic core dynamics of zinc finger miniproteins.

Philip Horx, Armin Geyer. Chem. Sci. 2021, 12, 11455-11463.

DOI: 10.1039/d1sc02346b

Supporting Information

Abstract The elucidation of internal dynamics in proteins is essential for the understanding of their stability and functionality. Breaking the symmetry of the degenerate rotation of the phenyl side chain provides additional structural information and allows a detailed description of the dynamics. Based on this concept, we propose a combination of synthetic and computational methods, to study the rotational mobility of the Phe ring in a sensitive zinc finger motif. The systematic methyl hopping around the phenylalanine ring yields o-, m-, p-tolyl and xylyl side chains that provide a vast array of additional NOE contacts, allowing the precise determination of the orientation of the aromatic ring. MD simulations and metadynamics complement these findings and facilitate the generation of free energy profiles for each derivative. Previous studies used a wide temperature window in combination with NMR spectroscopy to elucidate the side chain mobility of stable proteins. The zinc finger moiety exhibits a limited thermodynamic stability in a temperature range of only 40 K, making this approach impractical for this compound class. Therefore, we have developed a method that can be applied even to thermolabile systems and facilitates the detailed investigation of protein dynamics. 

Covalent linkage and macrocyclization preserve and enhance synergistic interactions in catalytic amyloids. 

Zsofia Lengyel-Zhand, Liam R. Marshall, Maximilian Jung, Megha Jayachandran, Min-Chul Kim, Austin Kriews, Ola V. Makhlynets, H. Christopher Fry, Armin Geyer, Ivan V. Korendovych. ChemBioChem 2021, 22, 585-591. 

DOI:10.1002/cbic.202000645

Supporting Informtion

Abstract How might complexity arise from short peptides? Synergistic interactions between the side chains of catalytic amyloids are locked into place by linkers between the peptides, and these new designs result in faster assembly and asymmetric assemblies. These could serve as a potential link in the evolutionary path between abiotic peptides and complex enzymes.

Tryptophan analogues with fixed side-chain orientation: expanding the scope. 

Lennart Nicke, Philip Horx, Ronny Müller, Sylvia Els-Heindl, Armin Geyer. ChemBioChem, 2021, 22, 330-335.

DOI: 10.1002/cbic.202000424

Supporting Information

Abstract A generalized synthetic strategy is proposed here for the synthesis of asymmetric β-indoylated amino acids by 8-aminoquinoline (8AQ)-directed C(sp3)-H functionalization of suitably protected precursors. Peptides containing one of the four stereoisomers of (indol-3-yl)-3-phenylalanine at position 2 of the parent peptide KwFwLL-NH2 (w=d-Trp) cover a wide range of activities as ghrelin receptor inverse agonists, among them the most active described until now. This application exemplarily shows how β-indoylated amino acids can be used for the systematic variation of the position of an indole group in a bioactive peptide.

Interactions of long-chain polyamines with silica studied by molecular dynamics simulations and solid-state NMR spectroscopy. 

Maria Montagna, Stephan Ingmar Brückner, Arezoo Dianat, Rafael Gutierrez, Fabian Daus, Armin Geyer, Eike Brunner, Gianaurelio Cuniberti. Langmuir 2020, 36, 11600-11609.

DOI: 10.1021/acs.langmuir.0c02157

Supporting Information

Abstract The investigation of molecular interactions between silica phases and organic components is crucial for elucidating the main steps involved in the biosilica mineralization process. In this respect, the structural characterization of the organic/inorganic interface is particularly useful for a deeper understanding of the dominant mechanisms of biomineralization. In this work, we have investigated the interaction of selectively 13C- and 15N-labeled atoms of organic long-chain polyamines (LCPAs) with 29Si-labeled atoms of a silica layer at the molecular level. In particular, silica/LCPA nanocomposites were analyzed by solid-state NMR spectroscopy in combination with all-atom molecular dynamics simulations. Solid-state NMR experiments allow the determination of 29Si–15N and 29Si–13C internuclear distances, providing the parameters for direct verification of atomistic simulations. Our results elucidate the relevant molecular conformations as well as the nature of the interaction between the LCPA and a silica substrate. Specifically, distances and second moments suggest a picture compatible with (i) LCPA completely embedded in the silica phase and (ii) the charged amino groups located in close vicinity of silanol groups.

(No graphical abstract available)

MON-185 ACTH-derived peptide antagonists as an alternative treatment strategy for congenital adrenal hyperplasia. 

J. Endocrin. Soc. 2020, 4, A298-A299. Tina Schubert, Lennart Nicke, André Schanze, Nicole Reisch, Armin Geyer, Katrin Koehler, Angela Huebner.

DOI: 10.1210/jendso/bvaa046.589

Abstract Congenital adrenal hyperplasia (CAH) is an autosomal recessive disorder caused by different enzyme deficiencies in the steroid hormone synthesis leading to a disturbed cortisol biosynthesis. The medical treatment of CAH includes suboptimal ACTH-suppressing high glucocorticoid doses to reduce adrenal hyperplasia and overproduction of androgens. These inappropriate corticoid substitutions are often associated with undesirable side effects such as arterial hypertension, growth failure and obesity. Since the current therapy of patients with CAH is often unsatisfactory, innovative treatment options are required.

The aim of our study was to specifically block the melanocortin 2 receptor (MC2R) signaling pathway as an alternative treatment strategy for CAH. We tested ACTH-derived selective peptides with incorporation of various synthetic non-natural amino acids in the activation motif of ACTH. To study the antagonistic activity of the peptides, cAMP production of MC2R/MRAP stably transfected human embryonic kidney (HEK) 293 cells were measured. All new synthesized peptide antagonists reduced ACTH-stimulated MC2R activity as competitive inhibitors indicated by a reduced in vitro cAMP response. Cells pre-incubated with peptide LNP009 showed the most efficient blockade of the MC2R and the highest shift of EC50 of ACTH (33.8 nM ± 0.08 vs. 7.3 nM ± 0.09). LNP009 was additionally tested for specificity concerning the other known melanocortin receptors and showed no antagonistic effect up to 1 μM on MC3, MC4 or MC5 receptor transiently transfected HEK 293 cells. To further investigate the inhibitory effect of our most potent antagonist peptide LNP009 on the steroid hormone response, we assessed steroidogenic enzyme expression of the human adrenocortical tumor cell line NCI-H295RA and performed mass spectrometry analyzes of steroids in the cell culture supernatant. Pre-incubation with LNP009 reduced the expression of the genes CYP21A2, CYP11B1 and HSD3B2 in NCI-H295RA cells and significantly reduced the synthesis of aldosterone (P=0.046; n=3), cortisol (P=0.020; n=3) and corticosterone (P=0.035; n=3).

With the successful blocking of the ACTH binding and signal transduction by our antagonistic peptides, we anticipate an alternative approach for optimizing the treatment of CAH patients lacking the side effects of the currently used ACTH-suppressing corticoid therapy.

Phosphate-silica interactions in diatom biosilica and synthetic composites studied by rotational echo double resonance (redor) nmr spectroscopy.

Langmuir 2020, 36, 4332-4338.Felicitas Kolbe, Fabian Daus, Armin Geyer, Eike Brunner. 

DOI: 10.1021/acs.langmuir.0c00336

Supporting information

Abstract Biosilica is a biogenic composite material produced by organisms like diatoms. Various biomolecules are tightly attached or incorporated into biosilica. Examples are special proteins termed silaffins and long-chain polyamines (LCPAs). Presumably, these biomolecules are involved in the biosilica formation process. Silaffins are highly phosphorylated zwitterions with LCPAs post-translationally attached to lysine residues. In the present work, we use distance-dependent solid-state NMR experiments, especially the 31P{29Si} Rotational Echo Double Resonance (REDOR) technique, to study the environment of phosphate moieties in biosilica and in vitro synthesized SiO2-based composites. In contrast to the heterogeneous mixtures of biomolecules found in native biosilica, the described in vitro silicification experiments make use of a single synthetic phosphopeptide and an LCPA of well-defined and uniform structure. The heteronuclear correlations measured from these silica composites provide reliable 31P–29Si dipolar second moments and information about the distribution of the phosphopeptide within the silica material. The calculated second moment indicates close contact between phosphopeptides and silica. The phosphopeptides are incorporated into the silica composite in a disperse manner. Moreover, the REDOR data acquired for diatom biosilica also imply that phosphate groups are part of the silica–organic interface in this material.