The Homunculus and the Paracelsian Liber de imaginibus

As William R. Newman has already shown, the alchemical homunculus described in the pseudo-Paracelsian writing De natura rerum was not the only kind of “homunculus” present in the works of (or attributed to) Paracelsus. Two other important kinds of “homunculi” indeed appeared in other treatises: one in De homunculis et monstris and the other in both Vom langen Leben and the Liber de imaginibus. This article focuses on the latter tract and its relationships with De natura rerum. After discussing the authenticity of the Liber de imaginibus, I will provide a brief analysis of its content and discuss the major topics common to the two treatises: the “signatures of things” and the homunculus. By studying the reception of the latter, I will show how the alchemical conception of the homunculus, as explained in De natura rerum, quickly established itself as the most prominent notion despite the fact that the golem-like version of Vom langen Leben and De imaginibus had nearly as much success at first among Paracelsians.     

A New Orthogonal Suppressor tRNA/Aminoacyl‐tRNA Synthetase Pair for Evolving an Organism with an Expanded Genetic Code

Several steps have been completed toward the development of a method for the site‐specific incorporation of unnatural amino acids into proteins in vivo. Our approach consists of the generation of amber suppressor tRNA/aminoacyl‐tRNA synthetase pairs that are orthogonal to all Escherichia coli endogenous tRNA/synthetase pairs, followed by directed evolution of the orthogonal aminoacyl‐tRNA synthetases to alter their amino‐acid specificities. A new orthogonal suppressor tRNA/aminoacyl‐tRNA synthetase pair in E. coli has been derived from the Saccharomyces cerevisiae tRNA^Asp and aspartyl‐tRNA synthetase, and the in vitro and in vivo characteristics of this pair were determined. Two different antibiotic resistance selections were compared using this novel pair in an effort to develop a tunable positive selection for a mutant synthetase capable of charging its cognate suppressor tRNA with an unnatural amino acid.     

New Light on the Alchemical Writings of Michael Sendivogius (1566–1636)

The Polish alchemist Michael Sendivogius is best known for the influential Novum lumen chymicum, a work composed of three separate texts. Sendivogius's authorship was questioned in the mid-seventeenth century, and these reservations are still held by some modern historians. On the other hand, other early modern and modern readers not only accepted his authorship of all three texts, but also ascribed as many as eleven texts to him. This paper discusses the key works published under the anagrammatised name of Sendivogius with the aim of resolving the authorship question. Newly discovered evidence makes it possible to trace the circumstances leading to the publication of these works in much greater detail than previously, and to present new arguments affirming Sendivogius's authorship. In the Tractatus de sulphure, Sendivogius promised to write another two treatises, which readers sought to identify and (in some cases) to write themselves. This paper sets out, and rejects, the arguments for including them in the corpus of genuine Sendivogian writings.     

Total Synthesis and Biological Evaluation of the Antibiotic Lysocin E and Its Enantiomeric,Epimeric, and N‐Demethylated Analogues

Lysocin E, a macrocyclic peptide, exhibits potent antibacterial activity against methicillin‐resistant Staphylococcus aureus (MRSA) through a novel mechanism. The first total synthesis of lysocin E was achieved by applying a full solid‐phase strategy. The developed approach also provides rapid access to the enantiomeric, epimeric, and N‐demethylated analogues of lysocin E. Significantly, the antibacterial activity of the unnatural enantiomer was comparable to that of the natural isomer, suggesting the absence of chiral recognition in its mode of action.     

Total Synthesis and Biological Evaluation of the Antibiotic Lysocin E and Its Enantiomeric,Epimeric, and N‐Demethylated Analogues

Lysocin E, a macrocyclic peptide, exhibits potent antibacterial activity against methicillin‐resistant Staphylococcus aureus (MRSA) through a novel mechanism. The first total synthesis of lysocin E was achieved by applying a full solid‐phase strategy. The developed approach also provides rapid access to the enantiomeric, epimeric, and N‐demethylated analogues of lysocin E. Significantly, the antibacterial activity of the unnatural enantiomer was comparable to that of the natural isomer, suggesting the absence of chiral recognition in its mode of action.     

Cathepsin B‐Specific Metabolic Precursor for In Vivo Tumor‐Specific Fluorescence Imaging

Recently, metabolic glycoengineering with bioorthogonal click reactions has focused on improving the tumor targeting efficiency of nanoparticles as delivery vehicles for anticancer drugs or imaging agents. It is the key technique for developing tumor‐specific metabolic precursors that can generate unnatural glycans on the tumor‐cell surface. A cathepsin B‐specific cleavable substrate (KGRR) conjugated with triacetylated N‐azidoacetyl‐d ‐mannosamine (RR‐S‐Ac₃ManNAz) was developed to enable tumor cells to generate unnatural glycans that contain azide groups. The generation of azide groups on the tumor cell surface was exogenously and specifically controlled by the amount of RR‐S‐Ac₃ManNAz that was fed to target tumor cells. Moreover, unnatural glycans on the tumor cell surface were conjugated with near infrared fluorescence (NIRF) dye‐labeled molecules by a bioorthogonal click reaction in cell cultures and in tumor‐bearing mice. Therefore, our RR‐S‐Ac₃ManNAz is promising for research in tumor‐specific imaging or drug delivery.     

Synthesis of New Plant Growth Regulators,Isomeric 1-(Dimethyloctyl)- and 1-Alkylpiperidinium and -morpholinium Halides by Telomerization of Isoprene with Secondary Amines

The known plant growth regulator, Al'den [1-(3,7-dimethyloctyl)-1-allylpiperidinium bromide], and also 1-(3,7-dimethyloctyl)-1-methylpiperidinium iodide were synthesized by treatment with allyl bromide and methyl iodide, respectively, of exhaustively hydrogenated 1-nerylpiperidine which was obtained by anionic telomerization of isoprene with piperidine. Various quaternary ammonium salts having a terpene sub- stituent with unnatural dimethyloctane skeleton, which effectively stimulate florification of Aster Chinensis L., were prepared by the action of alkyl halides on telomerization products derived from isoprene and piperidine or morpholine in the presence of palladium complexes.     

Structural Properties of Hachimoji Nucleic Acids and Their Building Blocks: Comparison of Genetic Systems with Four,Six and Eight Alphabets

Expansion of the genetic alphabet is an ambitious goal. A recent breakthrough has led to the eight-base (hachimoji) genetics having canonical and unnatural bases. However, very little is known on the molecular-level features that facilitate the candidature of unnatural bases as genetic alphabets. Here we amalgamated DFT calculations and MD simulations to analyse the properties of the constituents of hachimoji DNA and RNA. DFT reveals the dominant syn conformation for isolated unnatural deoxyribonucleosides and at the 5′-end of oligonucleotides, although an anti/syn mixture is predicted at the nonterminal and 3′-terminal positions. However, isolated ribonucleotides prefer an anti/syn mixture, but mostly prefer anti conformation at the nonterminal positions. Further, the canonical base pairing combinations reveals significant strength, which may facilitate replication of hachimoji DNA. We also identify noncanonical base pairs that can better tolerate the substitution of unnatural pairs in RNA. Stacking strengths of 51 dimers reveals higher average stacking stabilization of dimers of hachimoji bases than canonical bases, which provides clues for choosing energetically stable sequences. A total of 14.4 μs MD simulations reveal the influence of solvent on the properties of hachimoji oligonucleotides and point to the likely fidelity of replication of hachimoji DNA. Our results pinpoint the features that explain the experimentally observed stability of hachimoji DNA.     

Enzymatic Synthesis of a DNA Triblock Copolymer that is Composed of Natural and Unnatural Nucleotides

DNA molecules have come under the spotlight as potential templates for the fabrication of nanoscale products, such as molecular‐scale electronic or photonic devices. Herein, we report an enhanced approach for the synthesis of oligoblock copolymer‐type DNA by using the Klenow fragment exonuclease minus of E. coli DNA polymerase I (KF^?) in a multi‐step reaction with natural and unnatural nucleotides. First, we confirmed the applicability of unnatural nucleotides with 7‐deaza‐nucleosides—which was expected because they were non‐metalized nucleotides—on the unique polymerization process known as the “strand‐slippage model”. Because the length of the DNA sequence could be controlled by tuning the reaction time, analogous to a living polymerization reaction on this process, stepwise polymerization provided DNA block copolymers with natural and unnatural bases. AFM images showed that this DNA block copolymer could be metalized sequence‐selectively. This approach could expand the utility of DNA as a template.     

Potent De Novo Macrocyclic Peptides That Inhibit O-GlcNAc Transferase through an Allosteric Mechanism

Glycosyltransferases are a superfamily of enzymes that are notoriously difficult to inhibit. Here we apply an mRNA display technology integrated with genetic code reprogramming, referred to as the RaPID (random non-standard peptides integrated discovery) system, to identify macrocyclic peptides with high binding affinities for O-GlcNAc transferase (OGT). These macrocycles inhibit OGT activity through an allosteric mechanism that is driven by their binding to the tetratricopeptide repeats of OGT. Saturation mutagenesis in a maturation screen using 39 amino acids, including 22 non-canonical residues, led to an improved unnatural macrocycle that is ≈40 times more potent than the parent compound (K_i^app=1.5 nM). Subsequent derivatization delivered a biotinylated derivative that enabled one-step affinity purification of OGT from complex samples. The high potency and novel mechanism of action of these OGT ligands should enable new approaches to elucidate the specificity and regulation of OGT.     

Directed Evolution of a Designer Enzyme Featuring an Unnatural Catalytic Amino Acid

The impressive rate accelerations that enzymes display in nature often result from boosting the inherent catalytic activities of side chains by their precise positioning inside a protein binding pocket. Such fine‐tuning is also possible for catalytic unnatural amino acids. Specifically, the directed evolution of a recently described designer enzyme, which utilizes an aniline side chain to promote a model hydrazone formation reaction, is reported. Consecutive rounds of directed evolution identified several mutations in the promiscuous binding pocket, in which the unnatural amino acid is embedded in the starting catalyst. When combined, these mutations boost the turnover frequency (k_cat) of the designer enzyme by almost 100‐fold. This results from strengthening the catalytic contribution of the unnatural amino acid, as the engineered designer enzymes outperform variants, in which the aniline side chain is replaced with a catalytically inactive tyrosine residue, by more than 200‐fold.     

Enzyme-catalyzed synthesis of natural and unnatural polysaccharides

Synthesis of natural and unnatural polysaccharide was achieved via “enzymatic polymerization” by utilizing a glycoside hydrolase as catalyst. Particularly, hyaluronan, chondroitin, and their derivatives belonging to glycosaminoglycans have been prepared using sugar oxazoline monomers designed on the basis of the concept “transition-state analogue substrate”. The oxazoline derivatives of N-acetylhyalobiuronate [GlcAβ(1→3)GlcNAc] and N-acetylchondrosine [GlcAβ(1→3)GalNAc], which have the repeating disaccharide structures of hyaluronan and chondroitin, respectively, were successfully polymerized by the catalysis of hyaluronidase, giving rise to synthetic hyaluronan and chondroitin. Their 2-substituted oxazoline derivatives were also polymerized to the corresponding N-acylated hyaluronan and chondroitin derivatives. Furthermore, N-acetylchondrosine oxazoline derivatives sulfated at the C4, the C6, and both the C4 and C6 of the GalNAc unit were catalyzed by hyaluronidase; the monomer sulfated at the C4 was polymerized to chondroitin 4-sulfate with well-defined structure, whereas the other two monomers were exclusively hydrolyzed to the corresponding disaccharides. These different kinds of natural and unnatural polysaccharides having relatively high molecular weights were produced in all cases by the catalysis of hyaluronidase. © 2006 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 44: 5014–5027, 2006     

New dsDNA‐Binding Hybrid Molecules Combining an Unnatural Peptide and an Intercalating Moiety

In our search for new dsDNA‐binding ligands, combinatorial chemistry was first applied to select unnatural oligopeptides with moderate affinity for dsDNA. To enhance the binding affinity of a heptapeptide lead structure, Ac‐Arg‐Ual‐Sar‐Chi‐Chi‐Tal‐Arg‐NH₂ (K_d=4.9?10^?4 M ), the compound was conjugated to different heteropolyaromatic moieties by means of a variety of linker arms. Glycine, β‐alanine, glycyl‐glycine, glycyl‐β‐alanine, γ‐aminobutyric acid, and 6‐aminocaproic acid were used as spacers, representing different lengths and/or flexibilities. The intercalators coupled to the oligopeptide were acridine, fluorenone, anthracene, anthraquinone, and 3,8‐diamino‐5‐methyl‐6‐phenylphenantridinium (methidium). The binding capacities of these new hybrid molecules to dsDNA have been investigated by gel retardation and footprinting assays. The results show that, by conjugating the unnatural oligopeptide to intercalators, the affinity for dsDNA could be enhanced more than 100‐fold. For methidium‐β‐alanyl‐glycyl‐Arg‐Ual‐Sar‐Chi‐Chi‐Tal‐Arg‐NH₂ (K_d of 2.1?10^?6 M ), the interaction with dsDNA was dominated by the intercalator in such a way that the sequence specificity of the heptapeptide was changed. The interaction with dsDNA of hybrid molecules of other intercalators was mainly governed by the oligopeptide, since the sequence selectivity of the heptapeptide was conserved. In general, the linker arm glycine (shortest spacer) and glycine‐β‐alanine were preferred over β‐alanine, glycyl‐glycine and the more‐flexible spacers γ‐aminobutyric acid and 6‐aminocaproic acid. This way new hybrid molecules endowed with dsDNA affinities of ca. 10^?6 M and displaying different sequence selectivities have been developed. Therefore, combinations of such unnatural peptides with intercalators can be used to broaden the knowledge about the sequence‐selective recognition of dsDNA.     

Ullmann‐Ma Reaction: Development,Scope and Applications in Organic Synthesis†

Copper‐catalyzed cross‐couplings of aryl halides and nucleophiles, traditionally called Ullmann‐type coupling reactions, were initially reported by Ullmann et al. from 1901—1929. A seminal report in 1998 by Ma et al. from Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences revealed an accelerating effect caused by amino acids, which brought Ullmann‐type coupling reactions into a ligand‐accelerating era. From 1999 to the first 10 years of 2000s, the first‐generation ligands were developed by many researchers and promoted Ullmann‐type coupling reactions of aryl iodides and bromides under relatively mild conditions. Amino acid ligands, developed by Ma and coworkers, are one class of the most important first‐generation ligands. In the second 10 years of 2000s, Ma et al. led the discovery of second‐generation ligands for copper‐catalyzed cross‐coupling reactions. Two great breakthroughs have been realized by using second‐generation oxalic diamide and related amide ligands, with aryl chlorides as general coupling partner and with low catalyst loadings. Now copper‐catalyzed cross coupling reactions of aryl halides and nucleophiles with amino acids or oxalic diamides and related amides as ligands are recognized as Ullmann‐Ma reactions and have found extensive applications in organic synthesis.     

Bispalladacycle‐Catalyzed Michael Addition of In Situ Formed Azlactones to Enones

The development and further evolution of the first catalytic asymmetric conjugate additions of azlactones as activated amino acid derivatives to enones is described. Whereas the first‐generation approach started from isolated azlactones, in the second‐generation approach the azlactones could be generated in situ starting from racemic N‐benzoylated amino acids. The third evolution stage could make use of racemic unprotected α‐amino acids to directly form highly enantioenriched and diastereomerically pure masked quaternary amino acid products bearing an additional tertiary stereocenter. The step‐economic transformations were accomplished by cooperative activation by using a robust planar chiral bis‐Pd catalyst, a Brønsted acid (HOAc or BzOH; Ac=acetyl, Bz=benzoyl), and a Brønsted base (NaOAc). In particular the second‐ and third‐generation approaches provide a rapid and divergent access to biologically interesting unnatural quaternary amino acid derivatives from inexpensive bulk chemicals. In that way highly enantioenriched acyclic α‐amino acids, α‐alkyl proline, and α‐alkyl pyroglutamic acid derivatives could be prepared in diastereomerically pure form. In addition, a unique way is presented to prepare diastereomerically pure bicyclic dipeptides in just two steps from unprotected tertiary α‐amino acids.