Yoshio Matsushima

Structure analyses of oligosaccharides by tagging of the reducing end sugars with a fluorescent compound.

Summary A potential aldehyde group of an oligosaccharide is combined with 2-aminopyridine by means of the reductive amination with sodium cyanoborohydride giving a fluorescent 2-aminopyridine derivative. The sugar derivative can be used for the purpose of the determination of 1, a degree of polymerization; 2, sequence of the sugar units; 3, the linkage points of the sugar units, and the preparation of finger prints.

Analysis of neutral sugars as glycamines using an amino acid analyzer.

Abstract A new method of the analysis of neutral sugars was developed based on the separation of their corresponding glycamines. Sugars and ammonia were combined by means of the reduction with sodlum cyanoborohydride. The glycamines thus obtained were quantitatively analyzed by an automatic amino acid analyzer. Satisfactery results were obtained in the analyses of the constituent sugars of several polysaccharides and glycoconjugates.

Demonstration of the Physiological Role of Autolysis by a Comparative Study with a Wild‐Type and its Non‐Autolytic Mutant of Micrococcus lysodeikticus (luteus) Cultivated with Externally Added Proteolytic Enzymes

The log phase cells of autolytic Micrococcus lysodeikticus (luteus) IFO 3333 did not autolyze when grown in the presence of trypsin although the growth curve and morphology of the cells were not influenced.

Cell Wall Autolysis in Log Phase Cells of Micrococcus lysodeikticus (luteus)

Log phase cells of Micrococcus lysodeikticus (luteus) IFO 3333 autolyzed when incubated at 37 C in 0.01 m sodium‐phosphate buffer pH 7.5. The enzyme involved in the autolysis was recovered mainly in an aqueous phase from cytoplasmic membranes and cytoplasmic materials treated with n‐butanol, and proved to be an N‐acetylmuramyl‐L‐alanine amidase. The autolysis of log phase cells suspended in autolyzing buffer was depressed by the addition of trypsin to the buffer.

Evidence for occurrence of a characteristic amino acid sequence of glycopeptides in the linkage region between peptide and carbohydrate

Summary The amino acid sequences of the several glycopeptides derived from the cuttlefish skin collagen were established and compared with those derived from vertebrate collagens. The result indicates that there exists the common amino acid sequence which seems to act as a recognition sign for the glycosyltransferase to attach the first sugar to the polypeptide chain.

Cell Wall-Bridge Maintaining Three Dimensional Structure of Cell Packets Formed by the Localized Suppression of Cell Separation of a Micrococcus lysodeikticus (luteus) Mutant

Cell packets of Micrococcus lysodeikticus (luteus) mutant strain MT grown in medium supplemented with trypsin consisted of a tetrad as the unit structure. An interstice was observed between the unit‐tetrads, and a three dimensional structure of cell packets was maintained by the cell wall‐bridge along the rim of the cell packets which linked each unit‐tetrad. This unique structure of strain MT cell packets seemed to occur when the cell separation was suppressed locally, i.e., when the cross wall inside the initial site of cell separation was cut off, while the wall outside the initial site of separation was not cut off but remained as a joint of the daughter cells.

Process of consecutive cell divisions and separations in a regular tetrads-forming mutant of Micrococcus lysodeikticus (luteus).

A mutant MT of Micrococcus lysodeikticus (luteus) IFO 3333, whose minimum growing unit is not a single cell, but a tetrad unlike the wild‐type divides by binary fission of each monococcus, and then separates first into two daughter tetrads, second into four tetrads and third into eight tetrads. The three planes of either the cell division or the cell separation are equivalent to one another and oriented at right angles in three dimensions, respectively. The process of consecutive cell divisions and separations of the mutant tetrads was schematically illustrated.

On the structure of galactosylhydroxylysine and glycopeptides derived from bovine tracheal cartilage

Abstract Galactosylhydroxylysine isolated from the alkaline hydrolysate of bovine tracheal cartilage was shown to be O-β-D-galactopyranosylhydroxylysine. The structures of several hydroxylysine-containing glycopeptides suggested the existence of a unique sequence around the glycosylated site with one exception in that an amino acid other than arginine was present in the third position from the glycosylated residue. Isolation of glycopeptides with an identical hexapeptide sequence yet with a different carbohydrate side chain suggested that the attachment of glucose to a polypeptide-bound galactose is not regulated by a short amino acid sequence in the glycopeptide region.

Chemical Evidence for the Phillips Model of the Action of Hen Egg White Lysozyme

A THREE-dimensional model of the ES-complex formed between a chito-oligosaccharide and hen egg white lyso-zyme was presented by Blake et al. as a result of X-ray crystallographic investigations1. We reported that partially O-carboxymethylated chitin, which had satisfactory solubility, was highly susceptible to the action of lysozyme2 where the contents of the carboxymethyl groups were very roughly 0.3 mole/mole of N-acetoglucosamine residue. It is to be expected that the fairly bulky O-substituents will not permit such a free binding as that between unsubstituted chito-oligosaccharides and the enzyme, but will force the enzyme to select some suitable portions of the substrate molecules for making the ES-complex. Although it is impossible to know the exact location of the O-substituents on the high molecular substrate, the location of those on the enzymatic reaction products—which are low molecular compounds—can be determined. The orientation of the O-substituents in the product molecules must be reflexions of the conformation of the binding sub-sites in the ES-complex, because those suitable portions of the substrate molecule must have been fitted in the sub-sites before being released as low molecular saccharides.

Site of Cellular Autolysis in Micrococcus lysodeikticus(luteus)as Seen by Electron Microscopy

The cellular autolysis of bacteria is a phenomenon attributable to the hydrolytic degradation of cell wall peptidoglycans by their own enzyme(s). Studies by light and electron microscopy on Bacillus cereus and Escherichia coli (9), Clostridium botulinum (6), Clostridium perfringens (5), Streptococcus faecalis (3, 4), group H streptococci strains Wicky and Challis (13), and Lactobacillus acidophilus (2) have revealed that cytoplasmic materials leak outward at the septal region by the autolysis, and that the autolysis was initiated at the septal site of dividing cells and at one pole of just separated cells, where cell wall materials were synthesized actively. In particular, Higgins et al (3) demonstrated by electron microscopy of thin sections that the initiation of cellular autolysis in dividing cells of S. faecalis was brought about by the dissolution of cell wall at the leading edge of nascent cross wall. In a previous work (12), we found that the cellular autolysis in log phase cells of Micrococcus lysodeikticus (luteus) IFO 3333 was due to hydrolysis of the amide linkages between the glycan and peptide moieties of peptidoglycans. In the present study, we have attempted to locate where the autolysis of M. lysodeikticus arises from, by electron microscopy of thin sections. It should be pointed out that M. lysodeikticus has three division planes definitely oriented in three dimensions as shown by the observation of Yamada et al (15) and Monodane et al (10, 11) by scanning electron microscopy, unlike the above mentioned bacterial strains which have only one division plane.