Tetsuya Tanino

Total synthesis of (-)-muraymycin D2 and its epimer.

Full details of the first total synthesis of (-)-muraymycin (MRY) D2 and its epimer, the antibacterial nucleoside natural product, are described. Key strategic elements of the approach include the preparation of the urea dipeptide moiety found in the muraymycins containing an L-epi-capreomycidine via a nitrene C-H insertion of the sulfamate 10 and the fully protected muraymycin skeleton at a late stage by an Ugi four-component reaction. Thus, the nitrene C-H insertion of the sulfamate 10 with 10 mol % of Rh(2)(esp)(2) catalyst gave the cyclic sulfamates 11a and 11b in 47% yield (11a:11b = 1:2.0). Construction of the cyclic guanidine skeleton was effected through the HgBr(2)-promoted cyclization of 42 followed by desulfonylation upon acetolysis of the oxathiazinane ring to give 43 in good yield. The amine obtained by selective removal of the Cbz group of the alcohol 44 was reacted with MeSC(=O)-L-Val-O-t-Bu (38) to provide 45, which was oxidized to the carboxylic acid 46. Reaction of 46, isonitrile 51, isovaleraldehyde, and 2,4-dimethoxybenzylamine furnished the desired Ugi products, the final deprotection of which successfully afforded (-)-MRY D2 and epi-MRY D2 (53) after HPLC separation of the diastereomers. This approach would afford ready access to a range of analogues simply by altering each component.

Structural insights into inhibition of lipid I production in bacterial cell wall synthesis

Antibiotic-resistant bacterial infection is a serious threat to public health. Peptidoglycan biosynthesis is a well-established target for antibiotic development. MraY (phospho-MurNAc-pentapeptide translocase) catalyses the first and an essential membrane step of peptidoglycan biosynthesis. It is considered a very promising target for the development of new antibiotics, as many naturally occurring nucleoside inhibitors with antibacterial activity target this enzyme. However, antibiotics targeting MraY have not been developed for clinical use, mainly owing to a lack of structural insight into inhibition of this enzyme. Here we present the crystal structure of MraY from Aquifex aeolicus (MraYAA) in complex with its naturally occurring inhibitor, muraymycin D2 (MD2). We show that after binding MD2, MraYAA undergoes remarkably large conformational rearrangements near the active site, which lead to the formation of a nucleoside-binding pocket and a peptide-binding site. MD2 binds the nucleoside-binding pocket like a two-pronged plug inserting into a socket. Further interactions it makes in the adjacent peptide-binding site anchor MD2 to and enhance its affinity for MraYAA. Surprisingly, MD2 does not interact with three acidic residues or the Mg2+ cofactor required for catalysis, suggesting that MD2 binds to MraYAA in a manner that overlaps with, but is distinct from, its natural substrate, UDP-MurNAc-pentapeptide. We have determined the principles of MD2 binding to MraYAA, including how it avoids the need for pyrophosphate and sugar moieties, which are essential features for substrate binding. The conformational plasticity of MraY could be the reason that it is the target of many structurally distinct inhibitors. These findings can inform the design of new inhibitors targeting MraY as well as its paralogues, WecA and TarO.

Synthesis of l-epi-Capreomycidine Derivatives via C–H Amination

The L-epi-capreomycidine (Cpm) derivatives were efficiently and stereoselectively synthesized via nitrene C-H insertion starting from a readily available D-Tyr. Design of a substrate that takes into account hydrogen bonding is a critical feature in order to achieve high selectivity. Our synthetic strategy could be a new access to epi-Cpm and its derivatives, which are found in several biologically active natural products.

A New Arylsulfate Sulfotransferase Involved in Liponucleoside Antibiotic Biosynthesis in Streptomycetes

Sulfotransferases are involved in a variety of physiological processes and typically use 3′-phosphoadenosine 5′-phosphosulfate (PAPS) as the sulfate donor substrate. In contrast, microbial arylsulfate sulfotransferases (ASSTs) are PAPS-independent and utilize arylsulfates as sulfate donors. Yet, their genuine acceptor substrates are unknown. In this study we demonstrate that Cpz4 from Streptomyces sp. MK730–62F2 is an ASST-type sulfotransferase responsible for the formation of sulfated liponucleoside antibiotics. Gene deletion mutants showed that cpz4 is required for the production of sulfated caprazamycin derivatives. Cloning, overproduction, and purification of Cpz4 resulted in a 58-kDa soluble protein. The enzyme catalyzed the transfer of a sulfate group from p-nitrophenol sulfate (Km 48.1 μm, kcat 0.14 s−1) and methyl umbelliferone sulfate (Km 34.5 μm, kcat 0.15 s−1) onto phenol (Km 25.9 and 29.7 mm, respectively). The Cpz4 reaction proceeds by a ping pong bi-bi mechanism. Several structural analogs of intermediates of the caprazamycin biosynthetic pathway were synthesized and tested as substrates of Cpz4. Des-N-methyl-acyl-caprazol was converted with highest efficiency 100 times faster than phenol. The fatty acyl side chain and the uridyl moiety seem to be important for substrate recognition by Cpz4. Liponucleosides, partially purified from various mutant strains, were readily sulfated by Cpz4 using p-nitrophenol sulfate. No product formation could be observed with PAPS as the donor substrate. Sequence homology of Cpz4 to the previously examined ASSTs is low. However, numerous orthologs are encoded in microbial genomes and represent interesting subjects for future investigations.

Expansion of Antibacterial Spectrum of Muraymycins toward Pseudomonas aeruginosa.

It is urgent to develop novel anti-Pseudomonas agents that should also be active against multidrug resistant P. aeruginosa. Expanding the antibacterial spectrum of muraymycins toward P. aeruginosa was investigated by the systematic structure-activity relationship study. It was revealed that two functional groups, a lipophilic side chain and a guanidino group, at the accessory moiety of muraymycins were important for the anti-Pseudomonas activity, and analogue 29 exhibited antibacterial activity against a range of P. aeruginosa strains with the minimum inhibitory concentration values of 4-8 μg/mL.

Synthetic study of muraymycins using Ugi-four component reaction

The synthetic study of muraymycins (MRYs), which have potent antibacterial activity, is described. The key elements of our approach include the synthesis of L-epicapreomycidine via a C-H amination reaction and a conversient assemblage to construct of the framework of muraymycins using Ugi-four component reaction. First, isonitrile 4 was prepared from uridine in 14 steps. The precursor of carboxylic acid component 15 was synthesised via the C-H amination reaction, formation of cyclic guanidine structure. Muraymycin D2 analogs were synthesized by a model Ugi-four component reaction.