Tetsuo Shiba

Naofumi Uozumi; Shusaku Yanagidani; Eiji Miyoshi; Yoshito Ihara; Takahiko Sakuma; Congxiao Gao; Tadashi Teshima; Shigeru Fujii; Tetsuo Shiba; Naoyuki Taniguchi

GDP-L-Fuc:N-acetyl-β-D-glucosaminide α1→6fucosyltransferase (α1-6FucT; EC 2.4.1.68), which catalyzes the transfer of fucose from GDP-Fuc to N-linked type complex glycopeptides, was purified from a Triton X-100 extract of porcine brain microsomes. The purification procedures included sequential affinity chromatographies on GlcNAcβ1-2Manα1-6(GlcNAcβ1-2Manα1-2)Manβ1-4GlcNAcβ1-4GlcNAc-Asn-Sepharose 4B and synthetic GDP-hexanolamine-Sepharose 4B columns. The enzyme was recovered in a 12% final yield with a 440,000-fold increase in specific activity. SDS-polyacrylamide gel electrophoresis of the purified enzyme gave a major band corresponding to an apparent molecular mass of 58 kDa. The α1-6FucT has 575 amino acids and no putative N-glycosylation sites. The cDNA was cloned in to pSVK3 and was then transiently transfected into COS-1 cells. α1-6FucT activity was found to be high in the transfected cells, as compared with non- or mock-transfected cells. Northern blotting analyses of rat adult tissues showed that α1-6FucT was highly expressed in brain. No sequence homology was found with other previously cloned fucosyltransferases, but the enzyme appears to be a type II transmembrane protein like the other glycosyltransferases.

Masahiro Imoto; Hiroyuki Yoshimura; Nobuki Sakaguchi; Shoichi Kusumoto; Tetsuo Shiba

The first total synthesis of E. coli lipid A (1) is described. The synthetic compound was identical with a natural specimen and exhibited the full endotoxic activity. It was thus conclusively proved by this chemical synthesis that lipid A is the active principle of bacterial endotoxin.

Masahiro Imoto; Shoichi Kusumoto; Tetsuo Shiba; H. Naoki; T. Iwashita; E.Th. Rietschel; Horst-Werner Wollenweber; C. Galanos; Otto Lüderitz

Abstract The structure of the lipid A component of E. coli lippopolysaccharide was determined by means of chemical and 2D-NMR methods unequivocally to be a glucosamine β(1′-6)-disaccharide 1,4′-diphosphate acylated at the two hydroxyl (positions C-3 and −3′) and the two amino groups.

Masahiro Imoto; Shoichi Kusumoto; Tetsuo Shiba; E.Th. Rietschel; C. Galanos; Otto Lüderitz

The chemical structure of E. coli lipid A was elucidated to be 2 by determination of the nature of the individual acyl groups bound to the two hydroxyl groups in positions 3,3′ and the two amino groups of the D-glucosamine disaccharide phosphate backbone.

Tetsuo Shiba; Hiroyuki Mizote; Takeo Kaneko; Teruo Nakajima; Kakimoto Yasuo; Isamu Sano

Abstract A new basic amino acid, hypusine, was isolated from the homogenate of bovine brain tissue by ion-exchange column chromatography. The structure of this amino acid was determined to be N 6 -4-amino-2-hydroxybutyl)-2,6-diaminohexanoic acid on the basis of its physical properties involving NMR and mass spectra, as well as chemical degradation including periodate oxidation and reduction with HI and P.

Tateaki Wakamiya; Yasuyuki Ueki; Tetsuo Shiba; Yasuji Kido; Yoshinobu Motoki

Abstract The structure of ancovenin, a new peptide inhibitor of angiotensin I converting enzyme, was determined to be a unique tricyclic peptide which comprises sixteen amino acid residues including dehydroalanine and three sulfide amino acids as unusual components.

Ulrich Zähringer; Buko Lindner; Ulrich Seydel; E.Th. Rietschel; H. Naoki; Frank M. Unger; Masahiro Imoto; Shoichi Kusumoto; Tetsuo Shiba

Abstract The structure of the oligosaccharide portion of an E.coli Re lipopolysaccharide was determined as α-KDO-(2→4)-α-KDO-(2→6)-β-GlcN-α(1→6)-α-GlcN, bisphosphorylated at positions 1 and 4′. Taking into account the previous determination of the acylation pattern of the GlcN disaccharide, the total structure of E.coli Re LPS was thus established.

Koichi Fukase; Manabu Kitazawa; Akihiko Sano; Kuniaki Shimbo; Hiroshi Fujita; Shingo Horimoto; Tateaki Wakamiya; Tetsuo Shiba

Abstract Total synthesis of a lanthionine peptide nisin was successfully achieved for the first time by application of new methods for preparations of dehydroalanine and lanthionine moieties, resulting in a confirmation of the proposed structure.

Masahiro Imoto; Naoto Kusunose; Yoshiki Matsuura; Shoichi Kusumoto; Tetsuo Shiba

Abstract α-Ketopyranosyl fluorides, 2 and 3 , of 4,5:7,8-di-O-isopropylidene KDO methyl and benzyl esters were prepared and shown to act as effective glycosyl donors. X-Ray structure analysis of 2 and 1 H NMR study established the boat (B 3,6 ) conformation of the di-O-isopropylidene derivatives of KDO and enabled the assignment of their anomeric configurations.

Yoshitaka Nagai; Kyoichi Akiyama; Kenji Suzuki; Shozo Kotani; Yoshiro Watanabe; Tsutomu Shimono; Tetsuo Shiba; Shoichi Kusumoto; Fusahiro Ikuta; Shigeki Takeda

Abstract Mycobacteria in the adjuvant used for induction of experimental autoimmune encephalomyelitis (EAE) in guinea pigs can be replaced by synthetic N -acetylmuramyl- l -alanyl- d -isoglutamine. A combination of synthetic encephalitogenic peptides and muramyl dipeptides induces EAE effectively at a dose on the microgram level. In this system, the synthetic heptapeptide, H-Trp-Gly-Ala-Glu-Gly-Gln-Arg-OH, with a sequence identical to those of residues 116 to 122 of the basic protein of human myelin, was the shortest peptide causing EAE. These compounds form a simple system which should be useful in studies on the mechanism of the cell-mediated autoimmune reaction.