Effects of propionate on the growth of Rhodopseudomonas spheroides S were examined in the presence of various carbon compounds. Propionate strongly inhibits the growth on the medium containing glucose as a carbon source under aerobic-dark conditions, but not under anaerobic-light conditions. This inhibition can be restored by the addition of acetate or NaHCO₃ to the culture medium. Pyruvate, acetoin and acetaldehyde are accumulated in the culture medium during the growth inhibition induced by the addition of propionate. Metabolic pathway in the bacterium is discussed.

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Reduced and cyanoethylated glutenin was fractionated into three fractions (F I, F II and F III) by gel filtration on Sephadex G-100 in 0.1M acetic acid. The molecular weight determination was made with these three fractions by sedimentation equilibrium in 6.5M guanidine hydrochloride containing 0.01M acetic acid. The molecular weight obtained was 44, 000 for F II, and 32, 000 for F III. FI showed a distribution of molecular weight due to the aggregation. The avarage molecular weight of F I was 52, 000, being 27, 000 at the meniscus and 98, 000 at the bottom. Thte estimation of molecular weight by SDS-PAGE^* gave overestimated values for glutenin polypeptdes, as was already reported for gliadin.¹⁾

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Two lytic enzymes capable of lysing Streptococcus mutans have been purified to give a single band on disc-gel electrophoresis, respectively. The M-1 and M-2 enzymes were both proved to be N-acetylmuramidases. However, these enzymes were entirely different on their enzymatic properties. The molecular weights were about 20, 000 and 11, 000 for M-1 and M-2 enzymes, respectively, The maximal lytic activity of M-1 enzyme was obtained at ionic strength 0.05, while lytic activity of M-2 enzyme did not change within the ionic strength range of 0 to 0.05. The M-1 enzyme constituted the majority of the total lytic activity against the cell walls of Streptococcus mutans BHT of cultured filtrate. The M-2 enzyme showed less specific lytic activity on the cell walls of Streptococcus mutans BHT than M-1 enzyme.

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The preparation and properties of immobilized aminoacylase on porous glass by covalent binding [Porous glass-CVB-aminoacylase] and the continuous enzymatic reactions using such preparations are described.
Two types of porous glass-CVB-aminoacylase were prepared. One was aminoacylase covalently bound to alkylaminosilane derivative of porous glass with glutaraldehyde as a coupling agent [Alkylamino-porous glass-CVB-aminoacylase], and the other was aminoacylase covalently bound to arylaminosilane derivative of porous glass with nitrous acid as a coupling agent [Arylamino-porous glass-CVB-aminoacylase]. The enzyme activities of such immobilized aminoacylases were 3.2_??_13.0 units/ml glass for the former and 1.9_??_6.8 units/ml glass for the latter. Especially, alkylamino porous glass-CVB-aminoacylase showed excellent stability at pH 6_??_9 and temperature below 50°C, and was able to be stored for more than six months without appreciable loss of the activity.
The continuous enzyme reaction using the alkylamino porous glass-CVB-aminoacylase packed in a column was operated for 54 days at 37°C, and the half-life of the immobilized enzyme was calculated to be 78 days. From these results, it was recognized that such an immobilized aminoacylase on porous glass would be applicable in an industrial preparation of various L-amino acids from their DL-forms.

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This paper presents some data on general properties of the purified thiol: disulfide oxidoreductases (D-1-1) and (D-2) from Candida claussenii.
The optimum pH for both purified enzymes was pH 8.0 and the optimum temperatures for D-1-1 and D-2 were 40°C and 50°C, respectively. Both purified enzymes were stable below 30°C and in a pH range from 5.0 to 8.0 at 5°C. The enzyme activity was inhibited with several metals. Hg²⁺ and transition metals were the most toxic. The substrate specificities of both enzymes were wide, not only for the low molecular weight compounds but also for high molecular weight compounds, such as albumin and insulin. Various sulfhydrylgroups could serve as hydrogen donor.
Consequently, the reaction mode of these thiol: disulfide oxidoreductases was represented as follows:
R₁SSR₁+2R₂SH_??_2R₁SH+R₂SSR₂
Both enzymes had similar biochemical properties, but the molecular weights of D-1-1 and D-2 differed: the former, 155, 000_??_170, 000 and the latter, 29, 000_??_31, 000.
The enzyme activity was distributed in all subcellular fractions.

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Though sclerin (SCL) stimulated amino acid incorporation into the protein fraction of post mitochondrial supernatant of rat liver homogenate, it had no effect on the incorporation into the isolated mitochondria at pH 7.2, despite of its stimulating effect on mitochondrial oxidative phosphorylation. SCL stimulated amino acid incorporation into the mitochondria at pH 6.1, and to some extent maintained the activity on that in mitochondria during aging in hypotonic Tris-HCI buffer (pH 7.2). Since SCL prevented leakage of amino acids from the mitochondria into these buffers, it was suggested that SCL may protect a structure of mitochondria) membrane which appeared to have a significance on transport of amino acids. In liver slices, SCL stimulated amino acid incorporation only into the extra-mitochondrial fraction for the first 3min, but gradually turned to stimulate incorporation into mitochondria within 30min.

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A potent fungus for blood-coagulating enzyme was isolated from soil during screening tests of about 1500 strains of microorganisms. The fungus was identified as a strain belonging to Cephalosporium. This enzyme could be produced effectively by solid culture on wheat bran and partially purified to about 800 fold. This enzyme was one of the alkaline proteases. It was stable over a pH range of 7_??_10 and was activated by Ca²⁺ The enzyme could not coagulate 1% fibrinogen solution. The enzyme mixed with diluted blood plasma coagulated the fibrinogen solution. The enzyme also coagulated the citrated plasma from humans, bovines, mice, goats and hens.

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Isoleucyl-tRNA synthetase (IRS) was partially purified from an extreme thermophile, T. thermophilus HB 8. The molecular weight (11.5×10⁴) and some kinetic constants were obtained and compared with IRS from other sources.
The present IRS catalyzed both isoleucine dependent and valine dependent ATP-PP₁ exchange reactions (optimum at around 80°C) but not valyl-tRNA formation. The optimum temperature for isoleucyl-tRNA formation was 62°C with E. coli tRNA and 75°C with T. thermophilus tRNA.
The enzyme showed a remarkable thermostability. The addition of E. coli or T. thermophilus tRNA enhanced the thermostability of the enzyme, which was shown to be fully active up to 77°C. When E. coli tRNA was used, the loading activity decreased in parallel to the unfolding of the substrate tRNA molecule. From these results the relation is discussed between tRNA conformation and function.

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The structure of DNA extracted from dormant and germinating spores of B. cereus T was investigated using circular dichroism and other methods. No significant differences between DNAs extracted from vegetative cells and from spores of various stages could be found by analyses of CD spectra, CsCl density gradient centrifugation, melting profiles and template activity. All the DNA preparations were in B conformation and had the same density (1.695), Tm (83°C) and template activity in the reaction of DNA-dependent RNA polymerise. An abnormal DNA fraction of high density which was formerly found in B. cereus spores or a stable DNA complex with protein and/or RNA was not detected in the present extracts of spores. Preliminary X-ray analyses of intact spores indicate that the structure of DNA in spores is not so different from B form.

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Sclerin (SCL) stimulated the production of aminoglycoside antibiotics. Production of kanamycin (KM) was preceded by formation of KM-N-acetyltransferase and initiated by induction of N-acetyl-KM-amidohydrolase. KM-acetyltransferase rapidly developed and suddenly decreased at an early trophophase, whereas N-acetyl-KM-amidohydrolase appeared late and increased gradually. Addition of SCL to the culture initially most enhanced the productivity of KM, inducing both enzymes. Production of ribostamycin (RM) was also preceded by RM-acetyltransferase and optimal period for SCL addition was initial. On the other hand, production of streptomycin (SM) associated with both SM-(streptidino) kinase and alkaline phosphatase through trophophase and idiophase was rather stimulated by SCL added later. SCL induced alkaline phosphatase but not SM-(streptidino) kinase. Thus, a difference has been found in the effect of SCL between regulation of aminoglycoside-modifying (salvaging) enzymes and productivity of antibiotics.

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Quinolinate phosphoribosyltransferase has an important role in the NAD de novo biosynthetic pathway. Crystalline quinolinate phosphoribosyltransferase could be obtained for the first time from mammalian tissue. The crystalline enzyme preparation was certified to be homogeneous by polyacrylamide gel disc electrophoresis. Catalytic properties of this enzyme preparation were investigated. Optimum pH for the reaction was 6.1. Divalent cations were absolutely required and Mg²⁺ was the most effective. Michaelis constants for quinolinic acid and PRPP were 1.2×10^-4M and 1.8×10^-4M, respectively. Quinolinic acid could not be replaced by nicotinic acid or 2-amino nicotinic acid in this reaction. Di- and tri-valent cations fairly inhibited the reaction, but mono-valent cations had no effects. The reaction product was identified as β-nicotinic acid mononucleotide by its ultraviolet absorption spectra, paper chromatography, paper electrophoresis and its ORD spectrum.

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S-1358 was rapidly absorbed, metabolized and readily excreted via urine and feces from orally dosed rats. Excretion of radioactivity was almost complete within 4 days. The radioactivity was distributed mainly in stomach, intestines, liver and kidneys. It seems that S-1358 and its metabolites do not persist in organs and tissues following a single oral dosing.
Major urinary metabolites of the benzyl-labeled S-1358 were p-(1, 1-dimethyl-2-hydro-xyethyl) benzyl methyl sulfide [B], p-(1, 1-dimethyl-2-hydroxyethyl) benzyl methyl sulfone [A], p-(1-methyl-l-carboxylethyl) benzyl methyl sulfide [D], p-(1-methyl-l-carboxylethyl) benzyl methyl sulfone [C] and their glucuronide conjugates. Fecal metabolites were S-n-butyl S'-(l, 1-dimethyl-2-hydroxyethyl) benzyl N-3-pyridyldithiocarbonimidate [MR], A, B, C and D. These metabolites were also found in the bile. The pyridine-labeled S-1358 gave rise to 2-(3'-pyridylimino)-4-carboxylthiazolidine [HM] and 3-aminopyridine [AP] in the urine, and MR and AP in the feces. Intact S-1358 was a major component of the fecal radioactivity.

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3Z-Nonenal and 3Z, 6Z-nonadienal, potential biosynthetic precursors of 2E-nonenal and 2E, 6Z-nonadienal, were for the first time synthesized stereoselectively.

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Cytosine deaminase was purified about 900-fold from the cell-free extract of Serratia marcescens. The purification procedure included heat treatment, ammonium sulfate fractionation, ethyl alcohol fractionation, DEAF-cellulose and hydroxylapatite column chromatography, and Sephadex G-200 gel filtration.
The enzyme was homogeneous by the criteria of ultracentrifugation and acrylamide gel electrophoresis. The molecular weight was determined to be approximately 580, 000 and the molecule was composed of equimolecular weight of 8 subunits.
The enzyme catalyzed the stoichiometric conversion of cytosine into uracil and ammonia.

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Some physical, chemical and physiological properties of two S-CM subunits (S-CM IIe and S-CM Ala chain) of ricin D were studied. Physical properties of subunits are summerized in Table I. The S-CM subunits consisted of 244 (S-CM Ile chain) and 254 (S-CM Ala chain) amino acid residues. By the specific cleavage of the single intermolecular disulfide bond of ricin D, no remarkable change in conformation of polypeptide chains of ricin D was detected, but the toxicity was markedly decreased. The toxicity of ricin D is due to the quaternary structure of the ricin D molecule.

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The re-formation of the single disulfide bond which linked two polypeptide chains of ricin D was studied. Ricin D was reoxidized preferentially by the air-oxidation of its reduced polypeptide chains in a yield of 74% with the recovery of full toxicity. Furthermore ricin D was completely regenerated from its compound with p-chloromercuribenzoate. It seems reasonable to assume that the toxicity of ricin D arises from the quaternary structure of ricin D molecule.

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Ricin D, a toxic protein from castor bean, was found to contain 6 moles of glucosamine and 17 moles of mannose per mole of protein.
Isolation of two constituent polypeptide chains, namely Ala-chain and Ile-chain, and subsequent proteolytic digestions with Nagarse and Pronase revealed two glycopeptides (Asp₁, Thr₁ Gly₁, glucosamine₂ mannose₆ and Asp₁ Thr₁ Glu₁ Pro₁ glucosaminez mannose₇) from Alachain and one (Asp₁ Ile₁ Phe₁ glucosamine₂ mannose₄) from Ile-chain. The total carbohydrate content of these glycopeptides accounts for all that of the whole protein. It is therefore that carbohydrate moieties are covalently linked to the polypeptide chains in three sites to form this glycoprotein.

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Determination of the amino acid sequences of the three glycopeptides obtained from proteolytic digests of ricin D reveals that the site of carbohydrate linkage is at the Asx residue in all three peptides. Established structures of the glycopeptides are as follows:
Ala chain: (glucosamine₂ mannose₆)
Asx-Asx-Gly-Thr
(glucosamine₂ mannose₇)
Asx-Asx-Thr-Glu-Pro
Ile Chain: (glucosamine₂ mannose₄)
Ile-Asx-Phe

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Penicillin acylase (EC 3. 5. 1. 11) of Kluyvera citrophila KY7844 was purified approximately 120-fold by DEAE-cellulose chromatography, hydroxyapatite chromatography and isoelectrofocusing fractionation. The purified enzyme, with an approximate molecular weight of 63, 000, appeared to be homogeneous in disc electrophoretic analysis, and showed isoelectric point (Ip) 8.12 and 13.0 units/mg of specific activity for cephalexin hydrolysis. The Michaelis constant (Km) for cephalexin and for 7-[1-(1H)-tetrazolylacetamido]-desacetoxycephalosporanic acid ((1H) T-7ADCA) was 1.4mm and 3.6mm, respectively. This enzyme was capable of producing (1H) T-7ADCA in 80% yield from 1-(1H)-tetrazolylacetate methylester and 7-aminodesaceto-xycephalosporanic acid.

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The hydrolysis of acetal derivatives of oxidized spermine, N, N'-bis (3, 3-diethoxypropyl)-1, 4-diaminobutane, and its N, N'-dimethyl analog with acid was practically complete within 1 hr. During the hydrolysis of these compounds, no detectable amounts of acrolein were formed in the reaction mixture. While the hydrolyzate of the acetal derivative of oxidized spermine potently inactivated bacteriophage T₁, that of the N, N'-dimethyl analog had little phagocidal activity.

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A new method which utilizes ¹³C-¹³C coupling for structuual and biosynthetic studies on acetate-derived metabolites is describde. The ¹³C-NMR spectra of dihydrolatumcidins separately labeled with ¹³CH₃¹³CO₂Na and a 1:1 mixture of ¹³CH₃CO₂Na and CH₃¹³CO₂Na gave enough information to establish its structure.

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