Transketolase (TK, EC 2.2.1.1) was purified from bakers' yeast and spinach leaves by immunoaffinity using rabbit polyclonal anti-TK antibodies, which had been purified by immunoaffinity on immobilized TK, and coupled to Trisacryl. This enzyme is a useful tool for the synthesis of chiral ketoses. The properties of the enzymes from both origins were compared.

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The flow behavior and dynamic viscoelasticity of welan gum solutions were measured with a rheogoniometer. The welan gum showed shear-thinning behavior at a concentration of 0.1%, but plastic behavior above 0.3% at 25°C. The dynamic viscoelasticity increased with increasing concentration, and was scarcely changeable with increasing temperature even at 80°C. Gelation did not occur even in a polysaccharide concentration of 1.0% at low temperature (0°C). An increase of the dynamic modulus was not observed on the addition of CaCl₂ (6.8 mM). The dynamic viscoelasticity of welan gum solution was scarcely changeable in a wide range of pH from 2 to 12. The dynamic modulus was also scarcely changeable on addition of urea (4.0 M). Possible mode of intramolecular associations between the OH-4 of the D-glucosyl residue and the adjacent hemiacetal oxygen atom of the L-rhamnosyl residue, and between the methyl group of the L-rhamnosyl residue and the adjacent hemiacetal oxygen atom of the D-glucosyl residue were proposed.

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High hydrostatic pressures of 100 MPa to 300 MPa were applied to beef post-rigor muscle to investigate the efficiency of pressurization as a meat tenderizer.
The fragmentation of myofibrils increased with increasing pressure applied to the muscle, and the degree of fragmentation reached to its maximal level after briefly exposing (5 min) post-rigor muscle to the highest pressure (300 MPa). Electron microscopic studies of the pressurized muscle revealed that marked rupture of I-band and loss of M-line materials had progressed in the myofibrils with increasing applied pressure. However, degradation of the Z-line in myofibrils that can be observed naturally in conditioned muscle was not apparent in the pressurized muscle. There was no significant difference in the electrophoretic pattern of myofibrillar protein among the control and pressurized muscle samples in spite of the marked change of infrastructure.
From the results, it is suggested that the application of a high hydrostatic pressure to post-rigor muscle causes tenderization of the meat in a different manner from that of conditioning.

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2'-Deoxypuromycin (2) was synthesized to learn the effect of the 2'-hydroxyl group on the biological activity. Acylated xylose 3 was condensed with silylated 6-chloropurine to give β-D-xylofuranosyl-6-chloropurine derivative 4, whose 6-dimethylamination, 2'-deoxygenation and deprotection afforded 2'-deoxy-β-D-xylofuranosyl purine analog 7. The latter was converted to 2'-deoxypuromycin (2) in 8 steps. 2'-Deoxy analog 2 showed only weak antimicrobial activity compared with that of puromycin (1).

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The antioxidant activity of whole egg, egg albumen and egg yolk was estimated, and it was found that egg yolk had strong antioxidant activity on linoleate in an emulsion both with and without Fe²⁺. The relationship between the antioxidant activity and the components of the egg yolk was also investigated. The low-density lipoprotein (LDL) fraction of yolk had very weak activity, the granule fraction having the strong antioxidant activity in egg yolk. Phosvitin, a potent antioxidant, had weak activity in this system, but it was elevated by combining with the LDL fraction. When egg yolk was heated at 80°C for 30 min, its activity decreased. These results suggest that egg yolk had an antioxidant effect on linoleate in emulsions with and without Fe²⁺, and that granules, and not LDL, were the strong antioxidant fraction of egg yolk. Furthermore, it is suggested that both a phosvitin and native lipoprotein structure may be necessary for the antioxidant activity of the yolk granules.

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Light emission and the ESR spectra of tea leaves treated in various ways were studied. The fresh leaf showed a delayed light emission, which decreased exponentially with half times of 15 and 54 seconds. When the leaf was ground in water, the delayed light emission became small, because of destruction of the chloroplasts, and almost time-independent chemiluminescence was emitted. The chemiluminescence became very strong when the leaf was ground in an NaOH solution. ESR spectra of the tea leaf ground in water showed a singlet absorption attributable to an organic free radical, overlapping the six-line absorption due to the Mn²⁺ ion. The intensity of the radical became high when the leaf was treated with the NaOH solution and increased with time. In the case of the leaf ground in an NaCl solution, the singlet signal did not appear. These results suggest that the chemiluminescence and singlet absorption of the ESR spctra are attributable to oxidation of the catechins.

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A bovine β-Mactoglobulin (β-LG) was expressed in Saccharomyces cerevisiae carrying bovine pre-β-LG cDNA and secreted into its growth medium. The expression plasmid was constructed by inserting the whole coding region of the cDNA encoding pre-β-LG between the promoter and terminator of the yeast glyceraldehyde 3-phosphate dehydrogenase gene of pYG100, a yeast expression vector. In the supernatant of the yeast growth medium, β-LG with a native conformation was detected by sandwich ELISA, and its amount was estimated to be 1.1 mg/1. A Western-immunoblotting analysis revealed that β-LG secrected in the growth medium had the same mobility as that of authentic bovine β-LG. The N-terminal sequence was also identical with that of authentic mature bovine β-LG.

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When Hiproly barley callus was placed on an auxin-free Murashige and Skoog's medium, adventitious roots were formed. The contents of S-adenosyhnethionine (SAM) and 1-methylhistidine increased gradually in the callus throughout the root differentiation, along with increases in the levels of their precursors, methionine, histidine, and adenosine 5'-triphosphate. In the callus after auxin withdrawal, the addition of methionine resulted in rapid increases in the content of SAM and the incorporation of [methyl-¹⁴C]methionine into SAM. The accumulation of SAM was closely related to the elevation in methionine level in the callus. 1-Methylhistidine also increased greatly in the callus treated with methionine, but the addition of histidine had little effect on the change in 1-methylhistidine in the callus.

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Methanol dehydrogenase (MDH) and aldehyde dehydrogenase (ALDH) were purified to a homogenous state from Methylobacillus glycogenes, an obligate methylotroph. MDH (Mr 140, 000) was composed of two different subunits (M_r 60, 000 and 9, 000) forming an α₂/β₂ structure. MDH was indicated as a metalloquinoprotein containing one atom of calcium (Ca) per enzyme molecule. Binding of Ca was so tight that it was hard to remove Ca completely without denaturation of enzyme protein. A partially resolved enzyme resumed its original enzyme activity upon exogenous addition of Ca. Purified ALDH (M_r 144, 000) was composed of two identical subunits of molecular mass of 72, 000. ALDH was proved to be a quinoprotein in which PQQ is bound covalently.

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By applying the fractal concept, the relationship between particle diameter d_p and specific surface area S_w measured through an N₂ adsorption isotherm was investigated by using the equation S_w∝d^Ds-3_p where D_s is the fractal dimension to be in the range between 2 and 3. The surfaces of porous bodies of rice flour, wheat flour, corn fiber and milk powders were fractal, with D_s values between 2.26 and 3.0. The effect of defatting was also investigated.

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Transfructosylated derivatives of the steviol glycosides stevioside and rubusoside were synthesized by β-fructofuranosidase from Arthrobacter sp. K-1. The enzyme transferred a fructosyl residue regioselectively to the glucosyl moiety at the 19-carboxyl group of these steviol glycosides by β-2, 6-linkage. Evaluation of the sweetness of these fructosylated derivatives, in comparison with starting materials, found a great improvement in the quality of taste. Especially, the quality of taste of fructosyl-stevioside was superior to that of rebaudioside A, which is the best sweet principle in natural steviol glycosides in the intensity of sweetness as well as quality of the taste, being comparable to that of aspartame.

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Saccharomyces cerevisiae YNN27 (MATα. trp1 ura3 gal2) was mutagenized with ethyl methanesulfonate to isolate mutants deficient in glutathione biosynthesis. After mutants sensitive to 6.5 mM methylglyoxal were screened for, 12 glutathione biosynthesis-deficient mutants were isolated. Complementation tests found that the 12 mutants were classified into two different classes (YH1, YL1), and that each mutant was recessive and had a single mutation in glutathione biosynthesis. The growth of both strains in minimal medium was slower than that of the parent strain and was restored to the wild type level by supplementation with glutathione. They were also more sensitive to some chemicals such as thiol-reactive agents and metal-chelating agents than the parent strain. The mutant strain YL1 lacked activity of glutathione synthetase, while the deficiencies in γ-glutamylcysteine synthetase in mutant strains YH1 and YL1 were equivocal because the activity of the parent strain was very weak. However, supplementation with glutathione component amino acids and γ-glutamylcysteine to the medium showed that strain YH1, but not YL1, was deficient in γ-glutamylcysteine synthetase.

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Mutants resistant to the phenylalanine analogues o-fluoro-DL-phenylalanine (OFP) or p-fluoro-DL-phenylalanine (PFP) were isolated from a sake yeast, Saccharomyces cerevisiae Kyokai No. 9. They produced large amounts of β-phenylethyl alcohol and its acetate ester, which are rose-like flavor components. The tyrosine-dependent isozyme of 3-deoxy-D-arabino-heptulosonate-7-phosphate synthase (DAHP synthase; EC 4.1.2.15), in one of the OFP-resistant mutants was released from feedback inhibition by tyrosine. This mutant overproduced tyrosine in addition to β-phenylethyl alcohol and phenylalanine in minimal medium. In the PFP-resistant mutants, the activity of prephenate dehydrogenase (EC 1.3.1.12) was decreased to 5% of that in the wild type strain, while its DAHP synthase was increased four-fold. The tyrosine-dependent isozyme of this mutant was still sensitive to tyrosine. This mutant also overproduced tryptophan and phenylalanine in addition to β-phenylethyl alcohol in minimal medium.

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The sweet protein monellin consists of two noncovalently associated polypeptide chains, the A chain of 44 amino acid residues and the B chain of 50 residues. The [Asn²², Gln²⁵, Asn²⁶]-A chain, an analogue of the A chain, was synthesized by the stepwise Fmoc-solid-phase method in an overall yield of 13.4%. The synthetic A chain analogue was combined with the [Asn⁴⁹, Glu⁵⁰]-B chain, which was left over from a previous work, to give [Asn²², Gln²⁵, Asn²⁶]-A-chain-[Asn⁴⁹, Glu⁵⁰]-Bchain-monellin in a yield of 26.2%. This synthetic monellin analogue was approximately 550 times sweeter than sucrose. Changing the carboxyl groups of Asp²², Glu²⁵ and Asp²⁶ of the A chain to amide groups significantly decreased the sweetness potency. Crystallization was performed by a vapor diffusion method.

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We have identified by differential plaque hybridization, human cDNA clones encooding a member of a heat-shock protein family (hsp 90α) in the cDNA library of Adeno virus Type 12 ElA transfected HeLa cells. The complete nucleotide sequence of one of the clones (pHB76-114A) was identified. The sequence of 2912 base pairs had a single reading frame with a coding potential for an 84, 672-Da protein. The amino acid sequence was highly homologous, but not identical, to that of the human hsp 90a gene isolated from human peripheral blood lymphocytes [M. Yamazaki, K. Akaogi, T. Miwa, T. Imai, E. Soeda and K. Yokoyama, Nucleic Acids Res., 17, 7108 (1989)]. This cDNA hybridized with RNA species which increased 5- to 20-fold upon heat shock and more than 5-fold in the differentiation stage of human Tera 2 cells.

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The Retention Index (I) value and the different I values from two columns, carbowax 20M (20M) and OV101 (ΔI:I_20M-I_OV101) of aliphatic compounds were calculated by several kinds of linear equations under programmed linear temperature conditions. The equations could be used to predict appropriate compounds as well as a database of I and ΔI values.

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Isocitrate lyase was purified from the purple nonsulfur bacterium Rhodopseudomonas sp. No. 7. The purified enzyme was electrophoretically homogeneous. The molecular weights of the native enzyme and its subunit were estimated to be approximate 250, 000 and 62, 000 by gel filtration chromatography and SOS-polyacrylamide gel electrophoresis, respectively. The optimum pH for its activity was 6.5. The optimum temperature was 45°C. The Km for DL-isocitrate was 0.136 mM in potassium phosphate buffer (pH 6.0). Mg²⁺ was required for full activity of the enzyme as a non-essential activator. The enzyme activity was inhibited by SH-blocking reagents. Non-competitive inhibitory effects on the enzyme were examined with malate and succinate. The Ki for malate and succinate were 2.7 and 0.24 mM, respectively.

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A bacterium that assimilates 2, 3-dichloro-1-propanol was isolated from soil by enrichment culture. The strain was identified as Pseudomonas sp. by the taxonomic studies. The strain converted 2, 3-dichloro-1-propanol to 3-chloro-1, 2-propanediol, releasing chloride ion. The conversion was stereospecific because the residual 2, 3-dichloro-1-propanol and formed 3-chloro-1, 2-propanediol gave optical rotation. The resting cells converted various halohydrins to the dehalogenated alcohols, and cell-free extracts had strong epoxyhydrolase activity. These results indicated that the strain assimilated 2, 3-dichloro-1-propanol via 3-chloro-1, 2-propanediol, glycidol, and glycerol. The possibility to manufacture optically active 2, 3-dichloro-1-propanol is discussed.

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Oligosaccharides produced during the course of the hydrolysis of 25% N-acetylated chitosan by Streptomyces griseus chitinase were fractionated by CM-Sephadex C-25 and Toyopearl HW-40F column chromatographies. Sugar compositions and sequences of main oligosaccharides were identified by N-acetylation, exo-splitting with β-GlcNAcase and β-GlcNase, and nitrous acid degradation. In addition to N-acetylated saccharides, GlcNAc, (GlcNAc)₂, and (GlcNAc)₃, hetero-chitooligosac-charides such as GlcN•GlcNAc, GlcN• GlcNAc•GlcNAc, GlcN• GlcN•GlcNAc, GlcN• GlcNAc• GlcNAc• GlcNAc, GlcNAc• GlcN• GlcNAc• GlcNAc, GlcN• GlcNAc• GlcN•GlcNAc, and GlcN• GlcN •GlcNAc• GlcNAc were identified. These results indicate that Streptomyces griseus chitinase specifically cleaves the N-acetyl-β-D-glucosaminidic linkages in partially N-acetylated chitosan.

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The high immunogenecity of xylose-containing glycans of plant glycoproteins caused a non-specific antiserum which reacted to numerous glycoproteins of carrot cells and bromelain on immunoblots. The antiserum raised against a carrot M_r-57000 glycoprotein was fractionated by affinity chromatography, using a column in which bromelain was coupled with CNBr-Sepharose 4B. The passthrough and bound fractions contained anti-polypeptide moiety and anti-glycan moiety antibodies, respectively. The use of the pass-through fraction greatly improved the specificity when immunoblotting the total carrot proteins.

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Phosphodiesterase production with bis-p-nitrophenyl phosphate as a substrate by alkalophilic Bacillus No. A-40-2 increased with increasing Mn²⁺ concentration, showing maximum productivity at 10 mM. The enzyme production was negligible in the medium without Mn²⁺. The simultaneous addition of 10 mM Mn²⁺ and one of the several cations Mg²⁺, Co²⁺, Mo⁶⁺, and Pb²⁺ at suitable concentrations stimulated the enzyme production 1.8-fold at most over that with only 10 mM Mn²⁺ . Inorganic phosphate hardly repressed the enzyme production. The enzyme was purified homogeneously. The purified enzyme had the optimum pH of 7.5 and was fairly stable from pH 7-11. The enzyme hydrolyzed 2', 3'-cyclic-nucleotides and 3'-nucleotides, but did not hydrolyze 3', 5'-cyclic-nucleotides or 5'-nucleotides, indicating it to be a 2', 3'-cyclic-micleotide 2'-phosphodiesterase (EC 3.1.4.16). The enzyme had activity without metals, but Mg²⁺, Ca²⁺, Ba²⁺, and Mo⁶⁺ activated the enzyme reaction.

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Antihypertensive compounds were purified from an extract of autologous Lactobacillus casei cell lysates. The most effective compounds were polysaccharide-glycopeptide complexes, found in the cell wall. The average molecular weight was estimated as 180, 000 from gel filtration using Sephacryl S-300. The polysaccharide moiety of the complexes consisted of glucose, rhamnose, and galactose, whereas the glycopeptide moiety consisted of N-acetylglucosamine, N-acetylmuramic acid, asparagine, glutamine, alanine, and lysine. The varieties of the components of these moieties were constant and independent of complex molecular size. When these complexes were orally administered to spontaneously hypertensive rats (SHR) and renal hypertensive rats (RHR) at doses of 1 mg/kg-body weight, systolic blood pressure (SBP) decreased by 10-20 mmHg 6 to 12 hr after administration without any change in heart rate. Appreciable hypotensive activity was lost by treating the complexes with hydrofluoric acid, which hydrolytically cleaves the phosphodiester bond between the polysaccharide and glycopeptide moiety.

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A novel antibacterial compound, macrocarpal A, was isolated from the leaves of Eucalyptus macrocarpa, and its structure was determined on the basis of an X-ray crystal structure analysis. Macrocarpal A is composed of a phloroglucinol dialdehyde and diterpene, having a 3-membered ring, a 5-membered ring and a 7-membered ring.

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The gene coding for the ATCGAT specific BanIII DNA methyltransferase (M-BanIII) of Bacillus aneurinolyticus was cloned and its nucleotides sequenced. The coding region was assigned on the nucleotide sequence on the basis of the N-terminal amino acid sequence and molecular weight of the enzyme. The M-BanIII gene coded for a protein of 580 amino acid residues (MW 66, 344). Comparison with other methylases indicated that the M-BanIII sequence contained a segment of tetra-amino acids, NPPY, characteristic of N6-adenine methylases. In addition, some homologous regions were found in the sequences of type II adenine methylases PaeR7I(CTCGAG), TaqI((TCGA) and PstI(CTGCAG), containing TCGA within the recognition sequences.

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A enzyme that catalyzed the specific formation of ascorbic acid-2-phosphate (AsA2P) from ascorbic acid (AsA) and adenosine-5'-triphosphate (ATP), was purified 3, 200-fold to homogeneity from a cell extract of Pseudomonas azotocolligans. The purified enzyme appeared as a single band on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and consisted of a single polypeptide with a molecular weight of about 30, 000. Of phosphoryl donors tested, p-nitrophenylphosphate (p-NPP) and pyrophosphate (PPi) were as effective as ATP. Optimal pHs for the phosphorylating activity were around 4.0 and 5.5 when PPi and ATP were used as phosphoryl donors, respectively. The Km for AsA was 147 mM. The enzyme activity was inhibited by Cu²⁺, but not by sulfhydryl reagents.
The enzyme simultaneously had phosphatase activity at weakly acidic or neutral pH and the Km for p-NPP in the phosphatase activity was 0.38 mM. The enzyme was tentatively named "ascorbic acid phosphorylating enzyme."

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A DNA fragment of approximately 490 base pairs encoding human TNF was chemically synthesized and expressed within Escherichia coli cells. Furthermore, extracellular production of human TNF and several N-terminal deletion mutants of TNF was attempted using the excretion vector pEAP8. The TNF mutant with two N-terminal amino acids deleted (NΔ2-TNF) was efficiently excreted into the culture medium by E. coli carrying the plasmid pEXTNF3. In this clone, the signal peptide was correctly processed during the excretion. The E. coli-excreted NΔ2-TNF had higher antitumor activity than wild-type TNF or NΔ2-TNF produced intracellularly by E. coli.

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A variety of 3-O-[(3RS)-3-acyloxyacyl]-2-deoxy-2-[(3R)-3-hydroxytetradecanamido]-4-O-phosphono-D-glucose derivatives (41-52) carrying the acyloxyacyl groups of different carbon chain length at C-3 were synthesized from 2-(trimethylsilyl)ethyl 2-deoxy-4, 6-O-isopropylidene-2-[(3R)-3-[2-(trimethylsilyl)ethoxymethoxy]tetradecanamido]-β-D-glucopyranoside (1).

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The structures of sugar chains from the water-soluble glycoproteins in developing castor beans have been identified. The structural analyses were done by a fluorescence method combined with exoglycosidase digestions and ¹H-NMR spectroscopy. The identified structures fell into two categories; one was an oligomannose-type, the other xylomannose-type or xylose-containing type. Among these oligosaccharides, Man₃Fuc₁Xyl₁GlcNAc₂ (M3FX; 38%) and Man₆GlcNAc₂ (M6B; 22%) were the major structures. The higher mannose-content oligosaccharides (Man_8-7GlcNAc₂) were only 4.1%, and the further-modified structures (_GNM3FX, M2FX) than M3FX were 22% of the total.

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In organic solvents, lipase Amano P catalyzed the asymmetric ring opening of glutaric anhydrides bearing -CH₂Cl, -CH₂F and -OCH₃ groups at position 3 with methanol and 1-butanol to afford the corresponding mono esters in 70-86% e.e. Optically active δ-valerolactones were synthesized from the mono esters in overall yields of 34-62%. The -configuration was assigned to the mono esters and the lactones by means of a CD analysis and chemical correlation. This indicates that the lipase preferentially attacked the pro-R carbonyl group of these anhydrides carrying hetero atoms, in keeping with the trend observed for glutaric anhydrides having simple alkyl substituents.

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Although the growth of Brevibacterium flavum KH-21 with CS^L, PK^-, and PC^R was resistant to a lysine analogue, AEC, plus Thr at a concentration of 1 to 3 g/1, used conventionally for isolation of AK^R mutants, it was inhibited by higher concentrations of them, and recovered partially upon the addition of Lys, DAP and Met. The growth level recovered was equal to that in the presence of Thr alone. Several lysine-producing mutants with AK^R were isolated as those resistant to these high concentrations of AEC plus Thr. The representative strain, AH-198, produced 51 g/1 of Lys•HCl at maximum, when cultured for 72 hr in a medium containing 100 g/1 of glucose, while the parent strain KH-21 produced only 2 g/1. The productivity of strain AH-198 was the same as or a little higher than that of the HD^- type lysine-producing strain No. 22 with CS^L, PK^-, and PC^R. I_0.5, the concentration of Lys and Thr inhibiting 50% of the AK activity was 40 mM, 267-fold that of the parent enzyme. The Km for Asp and the optimum concentration of (NH₄)₂SO₄ for the apparent V_max were also increased. In addition, the inhibition by Thr or Lys alone disappeared in the mutant enzyme.

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A cell suspension culture of a Vitis hybrid converted quercetin to six glucosides. Their structures were identified as quercetin 3-O-β-D-glucopyranoside, quercetin 3, 4'-di-O-β-D-glucopyranoside, quercetin 3, 7-di-O-β-D-glucopyranoside, isorhamnetin 3-O-β-D-glucopyranoside, isorhamnetin 3, 4'-di-O-β-D-gIucopyranoside, and isorhamnetin 3, 7-di-O-β-D-glucopyranoside by UV, FD-MS, 1H-NMR, ¹³C-NMR spectroscopy and TLC analysis.
The course of conversion was also investigated and it was shown that quercetin 3-O-glucoside reached the maximum yield of 31% in 24hr and then gradually disappeared accompanied by the production of quercetin 3, 4'- and 3, 7-di-O-glucosides. Although the same rise and fall relationship was observed between isorhamnetin 3-O-glucoside and isorhamnetin 3, 4'- or 3, 7-di-O-glucoside, their conversion ratios were much lower than those of quercetin glucosides.

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The yeast Hansenula mrakii IFO 0895 could grow in a minimal medium containing 4 mM linoleic acid hydroperoxide, and the hydroperoxide moiety of linoleic acid hydroperoxide was reduced to the alcohol moiety. The activity of glutathione peroxidase was detected from the insoluble fractions of the cell homogenates of H. mrakii IFO 0895 only when the cells were grown in a linoleic acid hydroperoxide-containing medium.

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We examined the effects of tryptophan (Trp) on the hemagglutinating activity of Tora-mame (TM) lectin. Trp inhibited the hemagglutinating activity of TM lectin in a dose-dependent manner. On the other hand, other amino acids, i.e., alanine, phenylalanine, threonine, valine, methionine, arginine, leucine, and isoleucine, had no appreciable effect. Indoleacetic acid (IAA) also suppressed the hemagglutinating activity of TM lectin. The interaction of TM lectin with indole derivatives was also demonstrated by affinity chromatography with a Trp-coupled Sepharose 4B column. TM lectin was purified to apparent homogeneity from a crude extract of Tora-mame seeds by this affinity column chromatography. The hemagglutinating activities of other plant lectins, such as phytohemagglutinin (PHA), Ulex europeus lectin-I (UEA-I), Ricinus communis agglutinin (RCA), soybean lectin (SBA), peanut lectin (PNA), concanavalin A (ConA), wheat germ lectin (WGA), and Lentil lectin (LCA) were also modulated by Trp or IAA. These observations suggest that plant lectins may be important in cell elongation, division, and differentiation of higher plants through an interaction with indole derivatives.

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The complete amino acid sequence of antiviral protein from the seeds of pokeweed (Phytolacca americana) has been determined. This has been done by the sequence analysis of peptides derived by enzymatic digestion with trypsin, pepsin, and lysylendopeptidase, as well as by chemical cleavage with cyanogen bromide. The protein consists of 261 amino acid residues containing two disulfide bonds and has a calculated molecular mass of 29167 Da. The two disulfide bonds connect Cys-34 to Cys-258 and Cys-84 to Cys-105. Comparison of this sequence with the sequence of the ricin A-chain shows that there are identical residues at 76 positions in the two molecules (30% identity), having an extended region of highly conserved sequence at positions 170-183 (IQMXSEAARFXYIE). In contrast, the internal regions at positions 77-119 and 141-169 and the C-terminal 15 amino acid residues are less homologous in the two proteins.

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The restriction endonuclease AatII was purified from cell-free extracts of Acetobacter aceti IFO 3281 by streptomycin treatment, ammonium sulfate fractionation, combined column chromatographies on DEAE-Toyopearl 650S, heparin-Sepharose CL-6B and DEAE-Sepharose CL-6B and FPLC on Mono Q and on Superose 12 (gel filtration). The purified enzyme was homogeneous on SDSpolyacrylamide gel disk electrophoresis. The relative molecular mass of the purified enzyme was 190, 000 daltons by gel filtration. The SDS-polyacrylamide gel disk electrophoresis gave the relative molecular mass of 47, 500 daltons. These data indicated that the purified, native enzyme is a tetramer (190, 000 daltons) composed of four 47, 500-dalton subunits. The isoelectric point of the enzyme was 6.0. The purified enzyme was intensely activated by manganese ion (50-fold increase or more when compared with magnesium ion). The enzyme worked best at 37°C and pH 8.5 in a reaction mixture (50μl) containing 1.0μgλDNA, 10mM Tris-HCl, 7mM 2-mercaptoethanol, 7mM MnCl2 and 50mM NaCl. The enzyme recognizes the same palindromic hexanucleotide sequence 5'-GACGTC-3', cuts between T and C and produces a 3'-tetranucleotide extension in the presence of MnCl₂, as it does in the presence of MgCl₂.

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The monoacylation of (η⁶-1, 2-benzenedimethanol)tricarbonylchromium (2) by vinyl acetate, palmitate and benzoate, alcoholysis of the corresponding diesters of 2 in n-butanol, and acylation of (η⁶-benzyl alcohol) tricar bonylchromium by (±)-vinyl 2-phenoxypropanoate and 2-phenylpropanoate were accomplished with lipase P (from P. fluorescens) and lipase CC (from C. cylindracea) to give optically active organometallic esters. Their configurations indicated that the Stereoselectivity of each of these two Upases was in marked contrast. An active site model for them is proposed.

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