Hiroyuki Konno

Evaluation of peptide-aldehyde inhibitors using R188I mutant of SARS 3CL protease as a proteolysis-resistant mutant

n Abstractn n The 3C-like (3CL) protease of the severe acute respiratory syndrome (SARS) coronavirus is a key enzyme for the virus maturation. We found for the first time that the mature SARS 3CL protease is subject to degradation at 188Arg/189Gln. Replacing Arg with Ile at position 188 rendered the protease resistant to proteolysis. The R188I mutant digested a conserved undecapeptide substrate with a Kn m of 33.8μM and kn cat of 4753s−1. Compared with the value reported for the mature protease containing a C-terminal His-tag, the relative activity of the mutant was nearly 106. Novel peptide-aldehyde derivatives containing a side-chain-protected C-terminal Gln efficiently inhibited the catalytic activity of the R188I mutant. The results indicated for the first time that the tetrapeptide sequence is enough for inhibitory activities of peptide-aldehyde derivatives.n n

Synthesis of pyranicin and its inhibitory action with bovine heart mitochondrial complex I.

Total synthesis of pyranicin was achieved using Cl2Pd(CH3CN)2-catalyzed diastereoselective cyclization of the allylic ester as the key step. The inhibitory activity of this compound for mitochondrial NADH-ubiquinone oxidoreductase (complex I) was slightly poorer than that of ordinary mono-THF acetogenins such as cis-solamin.

Evaluation of lipid-binding properties of the N-terminal helical segments in human apolipoprotein A-I using fragment peptides

Although the N‐terminal region in human apolipoprotein (apo) A‐I is thought to stabilize the lipid‐free structure of the protein, its role in lipid binding is unknown. Using synthetic fragment peptides, we examined the lipid‐binding properties of the first 43 residues (1–43) of apoA‐I in comparison with residues 44–65 and 220–241, which have strong lipid affinity in the molecule. Circular dichroism measurements demonstrated that peptides corresponding to each segment have potential propensity to form α‐helical structure in trifluoroethanol. Spectroscopic and thermodynamic measurements revealed that apoA‐I (1–43) peptide has the strong ability to bind to lipid vesicles and to form α‐helical structure comparable to apoA‐I (220–241) peptide. Substitution of Tyr‐18 located at the center of the most hydrophobic region in residues 1–43 with a helix‐breaking proline resulted in the impaired lipid binding, indicating that the α‐helical structure in this region is required to trigger the lipid binding. In contrast, apoA‐I (44–65) peptide exhibited a lower propensity to form α‐helical structure upon binding to lipid, and apoA‐I (44–65/S55P) peptide exhibited diminished, but not completely impaired, lipid binding, suggesting that the central region of residues 44–65 is not pivotally involved in the formation of the α‐helical structure and lipid binding. These results indicate that the most N‐terminal region of apoA‐I molecule, residues 1–43, contributes to the lipid interaction of apoA‐I through the hydrophobic helical residues. Copyright

Thiamin-dependent transactivation activity of PDC2 in Saccharomyces cerevisiae.

MINT‐6797334, MINT‐6797355: THI3 (uniprotkb:Q07471) physically interacts (MI:0218) with PDC2 (uniprotkb:P32896) by two hybrid (MI:0018) MINT‐6797316: THI3 (uniprotkb:Q07471) physically interacts (MI:0218) with THI2 (uniprotkb:P38141) by two hybrid (MI:0018) MINT‐6797373: THI3 (uniprotkb:Q07471) binds (MI:0407) to PDC2 (uniprotkb:P32896) by pull down (MI:0096)

Ralstonins A and B, Lipopeptides with Chlamydospore-Inducing and Phytotoxic Activities from the Plant Pathogen Ralstonia solanacearum

Ralstonia solanacearum has an orphan hybrid polyketide synthase-nonribosomal peptide synthetase gene cluster. We herein isolate its products (named ralstonins A and B) from R. solanacearum and elucidate their structures and biological activities. Ralstonins are unusual lipodepsipeptides composed of 11 amino acids (containing unique amino acids such as β-hydroxytyrosine and dehydroalanine) and a 3-amino-2-hydroxyoctadecanoic acid, and their production is controlled by quorum sensing, a mechanism of bacterial cell-cell communication. Ralstonins exhibited chlamydospore-inducing activity and phytotoxicity.

Studies on Substrate Specificity at PR/p3 Cleavage Site of HTLV-1 Protease

Substrate specificities for recognition at the PR/p3 site of HTLV-1 protease were clarified using small libraries of substrate peptides. Specificities at P1 and P1′ positions were examined by parallel synthesis/digestion of synthetic peptides covering the PR/p3 site (KGPPVILPIQA). Specificities at P2 to P4 positions were examined by split and mix syntheses of olefin-peptide libraries containing the substrate sequence (PPVILPIQ). The solid-phase Horner-Emmons reaction was successfully applied to syntheses of multi-component substrates for library preparation. From the digestion of substrate peptides by a chemically synthesized mutant of HTLV-1 protease (C2A HTLV-1 PR), it was found for the first time that the preference for Pro at the P1′ position and for Ile at the P2 position is unique for this enzyme.

Synthesis of Squamostanal-A

The first synthesis of squamostanal-A (1), separated as a degradation product of tetrahydrofuranic acetogenins, is described. Iodide 7, which corresponds to the latent aldehyde moiety of 1, was prepared through a 2-step sequence from 13-[tetrahydropyran-2- yloxy]-2-tridecyn-l-ol (5). The NaHMDS-based coupling reaction of 7 with γ-lactone 8 gave compound 9, which by a 3-step sequence, was coverted to 1.