Tomoyuki Shibata

Documenting the scope of the catalytic asymmetric dihydroxylation

Abstract A wide variety of functionalized and unfunctionalized olefins are efficiently converted to the corresponding cis vicinal diols in moderate to good enantiomeric excess via the “slow addition” enhanced catalytic asymmetric osmylation process.

Synthetic study of 1-substituted carbapenem antibiotics

Abstract Total synthesis of 1-substituted carbapenems is described. The key step is the reaction of acetoxyazetidinone ( 7 ) with ketene silyl acetal ( 8 ).

A ligand structure-enantioselectivity relationship for the osmium-catalyzed asymmetric dihydroxylation of olefins

The highest enantioselectivities to date for various olefins are obtained in the osmium-catalyzed asymmetric dihydroxylation using new 9-O-aromatic dihydroquinidine and dihydroquinine ligands. A ligand structure-enantioselectivity relationship (LSER) is developed, and representative results for quinine-based ligands are reported for the first time.

Ligand-based improvement of enantioselectivity in the catalytic asymmetric dihydroxylation of dialkyl substituted olefins

A high level of asymmetric induction was achieved in the asymmetric dihydroxylation of dialkyl substituted olefins using 9-O-aryldihydroquinidines as ligands.

Identification of the biosynthetic gene cluster of A-500359s in Streptomyces griseus SANK60196.

A-500359s, produced by Streptomyces griseus SANK60196, are inhibitors of bacterial phospho-N-acetylmuramyl-pentapeptide translocase. They are composed of three distinct moieties: a 5′-carbamoyl uridine, an unsaturated hexuronic acid and an aminocaprolactam. Two contiguous cosmids covering a 65-kb region of DNA and encoding 38 open reading frames (ORFs) putatively involved in the biosynthesis of A-500359s were identified. Reverse transcriptase PCR showed that most of the 38 ORFs are highly expressed during A-500359s production, but mutants that do not produce A-500359s did not express these same ORFs. Furthermore, orf21, encoding a putative aminoglycoside 3′-phosphotransferase, was heterologously expressed in Escherichia coli and Streptomyces albus, yielding strains having selective resistance against A-500359B, suggesting that ORF21 phosphorylates the unsaturated hexuronic acid as a mechanism of self-resistance to A-500359s. In total, the data suggest that the cloned region is involved in the resistance, regulation and biosynthesis of A-500359s.

Functional and Kinetic Analysis of the Phosphotransferase CapP Conferring Selective Self-resistance to Capuramycin Antibiotics

Capuramycin-related compounds, including A-500359s and A-503083s, are nucleoside antibiotics that inhibit the enzyme bacterial translocase I involved in peptidoglycan cell wall biosynthesis. Within the biosynthetic gene cluster for the A-500359s exists a gene encoding a putative aminoglycoside 3-phosphotransferase that was previously demonstrated to be highly expressed during the production of A-500359s and confers selective resistance to capuramycins when expressed in heterologous hosts. A similar gene (capP) was identified within the biosynthetic gene cluster for the A-503083s, and CapP is now shown to similarly confer selective resistance to capuramycins. Recombinant CapP was produced and purified from Escherichia coli, and the function of CapP is established as an ATP-dependent capuramycin phosphotransferase that regio-specifically transfers the γ-phosphate to the 3″-hydroxyl of the unsaturated hexuronic acid moiety of A-503083 B. Kinetic analysis with the three major A-503083 congeners suggests that CapP preferentially phosphorylates A-503083s containing an aminocaprolactam moiety attached to the hexuronic acid, and bi-substrate kinetic analysis was consistent with CapP employing a sequential kinetic mechanism similar to most known aminoglycoside 3-phosphotransferases. The purified CapP product lost its antibiotic activity against Mycobacterium smegmatis, and this loss in bioactivity is primarily due to a 272-fold increase in the IC50 in the bacterial translocase I-catalyzed reaction. The results establish CapP-mediated phosphorylation as a mechanism of resistance to capuramycins and now set the stage to explore this strategy of resistance as a potential mechanism inherent to pathogens and provide the impetus for preparing second generation analogues as a preemptive strike to such resistance strategies.