Ryoji Yanagihara

Resolution of non-protein amino acids via microbial protease-catalyzed ester hydrolysis: marked enhancement of enantioselectivity by the use of esters with longer alkyl chains and at low temperature

Abstract In the microbial protease-catalyzed hydrolysis of amino acid esters with the free α-amino group, the enantioselectivity can be enhanced greatly by employing esters with longer alkyl chains such as the isobutyl ester instead of the conventional methyl ester and by conducting the reaction at low temperature.

Resolution of 2-aryloxy-1-propanols via lipase-catalyzed enantioselective acylation in organic media

Abstract 2-Aryloxy-1-propanols, primary alcohols with an oxygen atom at the stereocenter, were resolved with good to high enantioselectivity by acylation with vinyl butanoate mediated by Pseudomonas sp. lipase in di- iso -propyl ether. Potential factors affecting the enantioselectivity of the enzymatic acylation were examined: solvents, acyl donors and temperature. Using this enantioselective acylation procedure, enantiomerically pure ( R )-2-(4-chlorophenoxy)-1-propanol was prepared on a gram scale.

Remarkable effects of donor esters on the α-chymotrypsin-catalyzed couplings of inherently poor amino acid substrates

Abstract The extremely low efficiency during the α-chymotrypsin-catalyzed coupling of an inherently poor amino acid substrate, e.g. , alanine, using the methyl ester as an acyl donor was significantly improved using esters such as the 2,2,2-trifluoroethyl or carbamoylmethyl ester. The ameliorating effect of the latter ester was especially significant.

Resolution of 2-phenoxy-1-propanols by Pseudomonas sp. lipase-catalyzed highly enantioselective transesterification: influence of reaction conditions on the enantioselectivity toward primary alcohols

2-Phenoxy-1-propanols were resolved using Pseudomonas sp. lipase with E (enantiomeric ratio) values of ≥15. The enantioselectivity was highest with vinyl acetate and became lower with the chain length of the fatty acid moiety of acyl donors. It was improved with decreasing temperature. A linear direct correlation was observed between -RTlnE and the temperature of the reaction.

Broadening of the substrate tolerance of α-chymotrypsin by using the carbamoylmethyl ester as an acyl donor in kinetically controlled peptide synthesis

In the kinetically controlled approach of peptide synthesis mediated by α-chymotrypsin, the broadening of the protease’s substrate tolerance is achieved by switching the acyl donor from the conventional methyl ester to the carbamoylmethyl ester. Thus, as a typical example, the extremely low coupling efficiency obtained by employing the methyl ester of an inherently poor amino acid substrate, Ala, is significantly improved by the use of this particular ester. Its ameliorating effect is observed also in the couplings of other amino acid residues such as Gly and Ser as carboxy components.

A novel intramolecular hydrogen bonding between a side-chain pyridine ring and an amide hydrogen of the peptide backbone in tripeptides containing the new amino acid, α,α-di(2-pyridyl)glycine

Four tripeptides (Z-AA1-2Dpy-AA3-OMe; AA1, AA3 = Gly, Aib) containing a novel amino acid, α,α-di(2-pyridyl)glycine (2Dpy), were synthesized by the modified Ugi reaction. NMR analysis clearly indicated that the 2Dpy-containing tripeptides except the peptide in which AA1, AA3 = Aib, adopt a unique conformation with two intramolecular hydrogen bonds between 2Dpy-NH and a pyridine nitrogen and between AA3-NH and another pyridine nitrogen. This conformation has so far not been reported. On the other hand, the peptide Z-Aib-2Dpy-Aib-OMe probably adopts a β-turn structure which is stabilized by two intramolecular hydrogen bonds between 2Dpy-NH and a pyridine nitrogen and between AA3-NH and the CO of the Z group.

Conformational and coordination properties of a peptide containing the novel α,α-bis(2-pyridyl)glycine amino acidElectronic supplementary information (ESI) available: Figs. 1S, 2S. See http://www.rsc.org/suppdata/dt/b2/b209199b/

A fully protected tripeptide containing a novel Cα,α-disubstituted glycine—α,α-di(2-pyridyl)glycine (2Dpy)—in the central position and Cα,α-dimethylglycine (Aib) in positions 1 and 3 was synthesized and structurally characterized by single crystal X-ray analysis. The 2Dpy residue bears two 2-pyridyl moieties, which may coordinate metal ions, and promote self-assembly of peptide units. The tripeptide Z-Aib1-2Dpy2-Aib3-OCH3 (Z: benzyloxycarbonyl) adopts in the solid state a folded type III (III′) β-turn conformation (with Aib1-2Dpy2 as corner residues), stabilized by an i ← i + 3 hydrogen bond between the Aib3 NH and the urethane CO group. The peptide was able to self-assemble in the presence of Cu(II) ions, giving rise to an octahedral complex with a 2 : 1 peptide/Cu(II) stoichiometry, which has been structurally characterized. The metal ion, arranged on a crystallographic symmetry centre, is coordinated by two symmetry related 2Dpy residues, which act as tridentate ligands: each 2Dpy residue donates to the Cu(II) the N atoms of both pyridyl moieties in the equatorial positions, and the carbonyl oxygen atom in the axial positions. A very similar folded conformation characterizes the peptide in both the free and metal-bound states. This indicates that the overall peptide structure is ready to coordinate the metal ion without substantial conformational changes. Metal binding of the tri-peptide toward copper ions was analyzed by UV-visible spectroscopy performed in methanol solution. The experimental data agree with an equilibrium between two species in solution, one in the ratio Cu/peptide 1 : 1, the other 1 : 2. The formation constants are 9.7 × 103 M−1 and 3.5 × 102 M−1, respectively. The molar extinction coefficients are 66 ± 3 M−1 cm−1 (590 nm) for the 2 : 1 complex and 17 ± 2 M−1 cm−1 (690 nm) for the other. All the data obtained in the present work suggest that the 2Dpy residue is well suited to be accommodated into folded conformations. The spontaneous formation of an ordered structure by the 2Dpy containing tri-peptide and copper ions may guide toward the design of more elaborate self-assembling systems.

Superiority of the carbamoylmethyl ester as an acyl donor for the kinetically controlled amide-bond formation mediated by α-chymotrypsin

The superiority of the carbamoylmethyl ester as an acyl donor for the α-chymotrypsin-catalysed kinetically controlled peptide-bond formation is demonstrated in the couplings of an inherently poor amino acid substrate, Ala, with various amino acid residues as amino components and in the couplings of non-protein amino acids such as halogenophenylalanines as carboxylic components. Furthermore, this approach is applied to the amide-bond formation between an amino acid residue and a chiral amine, which is highly diastereoselective.

α-Chymotrypsin-catalysed peptide synthesis via the kinetically controlled approach using activated esters as acyl donors in organic solvents with low water content: incorporation of non-protein amino acids into peptides

The coupling efficiency in the α-chymotrypsin-catalysed peptide synthesis via the kinetically controlled approach is greatly improved by the use of activated esters such as the 2,2,2-trifluoroethyl ester as acyl donors instead of the conventional methyl ester in organic solvents such as acetonitrile with low water content. This approach is useful for the incorporation of non-protein amino acids such as halogenophenylalanines into peptides.

Enzymatic resolution of 2-phenoxy-1-propanols through the enantioselective acylation mediated by Achromobacter sp. lipase

Abstract2-(Substituted phenoxy)-1-propanols, e.g. 2-(4-chlorophenoxy)-1-propanol, belonging to primary alcohols with an oxygen atom at the stereocenter, were resolved with moderate to good enantioselectivity, as judged by the value of enantiomeric ratio E (up to 27), through the enantioselective acylation with vinyl butanoate mediated by the little-known lipase from Achromobacter sp. in diisopropyl ether, after the examination of potential factors affecting the reaction such as organic solvents and acyl donors.