Toshihiko Kanzaki

Accumulation of Tripeptide Derivatives by Mutants of Cephalosporium acremonium

The two β-lactam antibiotics produced by Cephalosporium acremonium ATCC 14553, cephalosporin C and penicillin N, have been proposed to be biosynthesized through the peptide, δ-(l-α-aminoadipyl)-l-cystemyl-d-valine.Many β-lactam negative mutants were derived from the strain No. 52, which was a more potent producer of the β-lactam antibiotics than the parent strain, C. acremonium ATCC 14553. Some of them were found to accumulate two sulfur-containing peptides. These compounds were isolated from the culture filtrate of one of the mutants, N-2 and determined to be the dimer of δ-(l-α-aminoadipyl)-l-cysteinyl-d-valine and the S-methylthio derivative of the tripeptide.

Studies on Streptomycetes

Investigation was made on the mycological and antibacterial properties of a strain No. 46408 isolated from a soil sample collected in Shimoueda, Wakayama Prefecture. The strain No. 46408 was compared with similar strains, St. hygroscopicus and St. halstedii, and it was judged to belong to a new species and therefore named Streptomyces atratus nov. sp. Also a strain A-165-Z1, which produces ilamycin, was referred to on its mycological properties and assumed to resemble St. atratus. St. atratus was found to produce new antibiotics, rufomycin A and B, specially active against acid-fast bacteria.

Biosynthesis of Cephalosporins

Publisher Summary This chapter discusses the biosynthesis of cephalosporins. A variety of cephalosporins and their producers has been found, however it has not been conclusively determined what the secondary metabolites are in a strict sense of the term. A moderate number of the metabolites are surmised in this chapter to be intermediates of the biosynthesis and degraded or modified products of secondary metabolites. It summarizes Cephalosporins and various metabolites related to the biosynthesis of β-lactam antibiotics in cephalosporin-producing strains. The features of the biosynthetic pathway of the β -lactam antibiotics, and especially of cephalosporins, have been disclosed through extensive work by several groups. The hypothesis that the tripeptide is a direct precursor of the β-lactam antibiotics was supported by the experiments of Fawcett et al. The most important problem about β-lactam antibiotic synthesis now involves the mechanism of the ring closure, i.e., the ability of the oxidation mechanism of the valine moiety to form the five-membered or the six-membered ring and of the cysteine moiety to form the β-lactam ring.

Quinones of Brevibacterium.

Abstract Two unusual menaquinones have been revealed during examination of Brevibacterium thiogenitalis and Brevibacterium vitarumen. Both microorganisms were found to contain menaquinones-8 (II-H2) and -9 (II-H2). The major quinone of the former was menaquinone-9 (II-H2) while that of the latter was menaquinone-8 (II-H2). No ubiquinones were demonstrated in both microorganisms.

l-Glutamic Acid Fermentation: Part VI. Structure of a Sugar Lipid Produced by Brevibacterium thiogenitalisPart VII. Relation between Biotin and Oleic Acid

Structure of a sugar lipid produced by an oleic acid-requiring mutant of Brevibacterium thiogenitalis was studied and established as (I).Relation between biotin and oleic acid was studied using a biotin-requiring organism accumulating l-glutamic acid and its blocked mutants lacking the biosynthetic system of biotin or/and oleic acid. The results support the following considerations. Biotin is not formed from oleic acid and does not substantially affect the growth of l-glutamic acid-accumulating bacteria and their productivity of l-glutamic acid.Consequently, biotin serves only for the synthesis of fatty acids in the present organisms. The essential factor for their growth and metabolism is an unsaturated fatty acid like oleic acid and not biotin. And also, saturated fatty acids have substantially no relation with their growth and metabolism like accumulation of l-glutamic acid.

L-Glutamic Acid Fermentation:Part V Behaviour of Oleic Acid in and Oleic Acid-Requiring Mutant

The effects of biotin and oleic acid concentrations in the media on the utilization and the distribution of oleic acid-U-14C were investigated using an oleic acid-requiring mutant of Brevibacterium thiogenitalis No. 653.Oleic acid in the media was rapidly taken up by the cells and its uptake-rate was enhanced by biotin. Oleic acid uptaken was incorporated into a sugar lipid and phospholipid fractions and some amounts of these lipids were excreted to the media.The distribution of oleic acid-U-14C in the cellular lipid fractions was changed by biotin or oleic acid concentration in the media. About 40% carbon of oleic acid which was added to the media was incorporated into the sugar lipid fraction.

Role of biotin in glutamate biosynthesis with a mixed substrate of glucose and acetate.

(1) Brevibacterium thiogenitalisから誘導されたオレイン酸要求株D-248をビオチン過剰(150μg/l以上)存在下で,グルコースと酢酸をモル比で1:2に混合して培養すると,グルタミン酸蓄積量が顕著に向上し,対炭素源当りの重量換算で実に70%の収率に達した.しかし,ビオチン添加量が少ないと,酢酸が残存し,ピルビン酸が著量蓄積した. (2) 乳酸やピルビン酸を炭素源とした培養でも,ビオチンとして100μg/l以上の添加が必要であり,さらに,休止細胞反応においても,ビオチンの添加効果が認められた. (3) 粗酵素液中に,ホスホエノールピルビン酸カルボキシラーゼ活性と同時にピルビン酸カルボキシラーゼ活性が検出された.ピルビン酸カルボキシラーゼ活性は,培地中に添加したビオチン量の増加に応答を示した. これらの結果より,ビオチンがピルビン酸カルボキシラーゼ活性を介して,グルタミン酸生合成に直接関与していることが明らかとなった.

l-Glutamic Acid Fermentation: Part III. The Physiological Characteristics of an Oleic Acid-Requiring MutantPart IV. The Relation between the Cellular Fatty Acid Contents and the Productivity of l-Glutamic Acid

A study was made on the differences between Brevibacterium thiogenitalis No. 653 and its oleic acid-requiring mutant D-248 in some physiological characteristics.The most important difference of the characteristics was found in their intracellular fatty acid contents. Namely, the cellular oleic acid content of D-248 was scarcely affected by biotin but limited by the oleic acid which was added to the medium.On the other hand, various enzyme activities and rates of oxygen uptake for several organic acids were found to be slightly different between the two strains.These observations suggest that oleic acid has an important role for the production of l-glutamic acid.The effect of biotin and oleic acid on the cellular fatty acid contents, and the relation between the cellular fatty acid contents and the productivity of l-glutamic acid were investigated using Brevibacterium thiogenitalis No. 653 and its oleic acid-requiring mutant, D-248.While the synthesis of palmitic acid in D-248 was stimulated by biotin and ...