Further characterization of phosphoinositol kinase isolated from germinating mung bean seeds

Phosphoinositol kinase isolated and purified from germinating mung bean seeds has been further characterized. The rate of phosphorylation varies with different inositol phosphates and this is consistent with the K_m and V_max for each of the substrates. The phosphate transfer from ATP has been found to be mediated by a phosphoprotein intermediate. In a particular step of the reaction the immediate product of the reaction has been found to be most inhibitory, other products being less or non-inhibitory. The inhibition has been found to be competitive in nature. The K_is have been found to range between 0.6 and 1 × 10^?4 M. ADP also inhibited non-competitively with respect to IP₅. K_i for this has been found to be 2.3 × 10^?4 M. The purified enzyme migrated as a single protein band on polyacrylamide gel electrophoresis. In the presence of sodium dodecyl sulphate it is dissociated into 3 subunits in the ratio 1 : 1 : 1. The MW of the three subunits are approx. 86 000, 56 000 and 35 000. The MW of the enzyme has been found to be approx. 177 000.     

Phosphoinositol kinase from germinating mung bean seeds

Phosphoinositol kinase (adenosine triphosphate-inositolmonophosphate phospho—tranferase) has been isolated from cotyledons; about 300-fold purification has been achieved, with a recovery of 11%. The enzyme has a pH optimum at 7·4. It can mediate phosphorylation of lower inositol phosphates to their corresponding higher homologues, ATP being the phosphate donor. ATP can be replaced partially by UTP and PEP. The enzyme requires divalent cations for the reaction. Mn²⁺ has been found to be twice as effective as Mg²⁺, Ca²⁺ being inhibitory. Phosphoinositol kinase has been found to be different from inositol kinase.     

Two forms of phosphoinositol kinase from germinating mung bean seeds

Phosphoinositol kinase, the key enzyme responsible for the biosynthesis of higher inositol phosphates has been isolated from the cotyledons of mung beans germinated for 24 hr and has been resolved into two different forms, phosphoinositol kinase A and phosphoinositol kinase B. Both forms were purified to homogeneity and characterized. The K_m values for ATP with phosphoinositol kinase A (1.78 × 10^?4 M) and phosphoinositol kinase B (3.12 × 10 ^?5 M) showed that phosphoinositol kinase B had a greater affinity for ATP. ATP could be partially replaced as phosphate donor by UTP and phosphoenolpyruvate in the case of phosphoinositol kinase A but not in the case of phosphoinositol kinase B.     

Changes in the forms of invertase during germination of mung bean seeds

High levels of ‘alkaline’ invertase activity occur in dormant mung bean seeds. During germination this activity decreases rapidly and is replaced by high ‘acid’ invertase activity. Cycloheximide prevented the formation of the latter activity and also inhibited germination. It is suggested that de novo synthesis of ‘acid’ invertase occurs during germination. Both enzymes bind to concanavalin A and, hence, are presumed to be glycoproteins. Affinity-purified enzyme samples show similar ratios of ‘acid’ and ‘alkaline’ invertase activities to the crude preparations indicating that specific enzyme inhibitors or activators are probably not involved in controlling the activities in vivo.     

Occurrence of glycoprotein glycosidases in mature seeds of mung bean (vigna radiata)

The presence of nine different glycosidases was demonstrated in the crude extract of mature mung bean seeds. N-Acetyl β-D-glucosaminidase, α-D-galactosidase and β-D-glucosidase were each resolved into two respective active forms by gel filtration. The other glycosidases showed single forms only. The apparent MWs of the glycosidases were determined. The glycosidases were absorbed to Con A-Sepharose column, with the exception of a small percentage of α-galactosidase and α-mannosidase which were eluted unretarded. The bound enzymes displayed varying affinities for the immobilized lectin, indicating differences in glycosylation. With the exception of β-galactosidase and invertase, all the glycosidase activities were detected in the protein bodies isolated from the seeds.     

Kinetic properties of IAA oxidase from mung bean cotyledons

A crude enzyme preparation from mung bean cotyledons was separated into peroxidative and non-peroxidative IAA oxidase on a DEAE-cellulose column. Both fractions differed in their pH optima, K_m and V_max. The K_m and V_max of non-peroxidative IAA oxidase were higher than those of peroxidative IAA oxidase. Peroxidative IAA oxidase showed a linear increase in absorption at 247 and 254 nm after a short lag of 2–3 min. The addition of catalytic amounts of hydrogen peroxide eliminated the lag period and also enhanced the rate of IAA degradation. The non-peroxidative IAA oxidase fraction, however, did not exhibit any significant increase in absorption at 247 and 254 nm and showed a lag period of 5 min which was not affected by hydrogen peroxide. Instead, addition of the same catalytic amount of hydrogen peroxide inhibited the rate of IAA degradation. The peroxidative IAA oxidase fraction exhibited the reaction kinetics characteristic of peroxidase-catalysed IAA degradation. The rate of IAA oxidation by purified non-peroxidative IAA oxidase was very low. The slow rate of catalysis shown by non-peroxidative IAA oxidase appears to be due to the presence of inhibitor(s).     

Regulation of sterol biosynthesis in etiolated mung bean hypocotyl sections

The incorporation of radioactivity into sterols by transmethylation of methionine-[¹⁴C-methyl] was studied in mung bean hypocotyl sections. Young hypocotyl sections (1 cm) synthesized 4 times more radioactive sterols than older sections (5 cm). The transmethylation reactions may be rate limiting in older tissues. Wounding has only a quantitative effect on sterol biosynthesis, as seen by incorporation experiments with MVA-[2-¹⁴C]. Naphthalene acetic acid (NAA) stimulates sterol biosynthesis in both wounded surfaces and intact tissues of mung bean hypocotyl sections.     

Indole plant-growth inhibitor from Abrus precatorius seeds

The structure of a new plant growth inhibitor isolated from the seeds of jequirity bean (Abrus precatorius) has been shown to be N,N-dimethyl-l-tryptophan.     

Coumaroyl-, caffeoyl- and feruloyltartronates and their accumulation in mung bean

Three new plant constituents were isolated from the primary leaves of Vigna radiata (= Phaseolus aureus) and their structures elucidated and characterized with the aid of negative-ion fast atom bombardment mass spectrometry (FAB MS), ¹H NMR and UV spectroscopy, thin-layer, gas-liquid and high performance liquid chromatography. The new conjugates are (E)-p-coumaroyl-, (E)-caffeoyl- and (E)-feruloyltartronic acids. Their structures were unequivocally confirmed by comparison with synthetic material. The metabolism of the new hydroxycinnamic acid conjugates in young plants of Vigna radiata is described.     

绿豆皮中黄酮类化合物提取工艺

研究了绿豆总黄酮的提取工艺。通过对固液比、乙醇体积分数、提取温度与提取时间的单因素实验确定水平点,设计4因素3水平实验,选用L9（3^4）正交表,优选绿豆总黄酮的最佳提取工艺为固液比1：50,φ（乙醇）为40％,提取温度70℃,提取时间120min。在此基础上得到绿豆皮中总黄酮的提取量为27．57mg／g,且5次平行的相对标准偏差为0．75％,总黄酮的平均回收率达97．5％。     

Effect of cations on the structure of mung bean cytoplasmic ribosomes

The importance of the absolute and relative concentrations of monovalent and divalent cations and centrifugal speed (pressure) in the dissociation of mung bean 80S ribosomes has been examined. In the absence of Mg²⁺ ions, ribosome monomers yield 47S and 34S particles. Fixation with glutaraldehyde, however, indicates that this dissociation pattern is largely dependent upon high pressures developed during centrifugation and that in the absence of such artifacts the immediate product of Mg-free conditions is a 74S particle. Since 74S particles rapidly revert to the 80S form when Mg is replaced, this would appear to be a conformational change. Ribosomes were also dissociated in the presence of Mg²⁺ ions if the K⁺ ion concentration was raised. Three major particles were produced, 38S and 49S from the small ribosomal sub-unit and 60S from the large sub-unit. A proportion of the 80S monomer population is more resistant to dissociation. Experiments with puromycin indicate that the more resistant fraction probably represents ribosomes completed with nascent polypeptide resulting from polysome breakdown.     

Absence of multiple forms of ribulose 15-bisphosphate carboxylase/oxygenase in mung bean

Ribulose 1,5-bisphosphate carboxylase/oxygenase has been reported to occur in multiple forms in mung bean (Phaseolus aureus) using Sephadex G-200 chromatography. We have isolated this enzyme by identical methodology. The profile from Sephadex G-200 chromatography shows only one peak in contrast to the previous report and we find no evidence to corroborate the conclusions. Where V_c, V_o and K_c, K_o represent V_max and Michaelis constants, respectively, the constant V_cK_o/V_oK_c for the single form is 70 at 40 μM CO₂ and 1200 μM O₂.     

Purification and partial characterization of an aminopeptidase from mung bean cotyledons

An aminopeptidase (EC 3.4.11.-) was purified to homogeneity, as judged by SDS-PAGE. from mung bean ( Vigna radiata ) cotyledons. The molecular mass of this peptidase was estimated as 75 kDa by gel filtration. When an oligopeptide consisting of 5 amino acid residues was used as substrate, amino acids were released in the order of the N-terminal sequence of the oligopeptide chain. This enzyme apparently requires free sulfhydryl for its activity, as judged by the effects of various proteinase inhibitors. Among aminoacyl- p -nitroanilides examined for the availability as substrates of the enzyme, p -nitroanilides with hydrophobic amino acids were preferred substrates. According to western immunoblot profiles, the enzyme level in cotyledons was high at the early stage of imbibition and declined rapidly after germination.     

低频电磁场与绿豆种子萌发

探讨低频电磁场的非热效应对绿豆种子萌发及其生长的影响,以及不同强度电磁场对种子萌发和生长的生物学效应。将绿豆种子置于低频电磁场(50Hz 2mT、4mT、6mT、8mT)下萌发、生长,对不同强度下种子萌发和生长情况进行观察分析表明：不同的电磁场强度对绿豆生物学效应是不同的;不同的电磁场强度对绿豆萌发以及随后的生长阶段的影响也是不同的。     

胃蛋白酶水解绿豆分离蛋白的工艺

选用胃蛋白酶对绿豆分离蛋白进行酶法水解,考察了原料预处理条件、pH、温度、底物浓度等对酶解的影响,结果表明：原料预处理最适条件为沸水浴中90℃处理20min,在37℃、pH1．8、底物质量分数7％、酶量6000U／g条件下酶解180min,水解度（DH％）为19．86％,达到了制备小肽的水解度要求。实验证明,经过水解,绿豆分离蛋白各功能特性得到很好的改善。     

Uncoupling activities of chalcones and dihydrochalcones on isolated mitochondria from potato tubers and mung bean hypocotyls

The uncoupling properties of 23 chalcones and dihydrochalcones were studied. Twelve compounds completely uncouple oxidative phosphorylation in mung bean hypocotyl and potato tuber mitochondria, four are weak uncouplers and seven are without effect. Usually, mung bean mitochondria are more sensitive to uncoupling agents than potato mitochondria. The uncoupling activity of chalcones and dihydrochalcones appears to be connected with the presence of hydrogen or hydroxyl groups in the 2′-position and hydrogen, hydroxyl or nitrate groups in the 4′-position. The α-β-unsaturated carbonyl system is not essential for activity. For the compounds which are not very lipophilic, substituents on the B-ring are without effect on the uncoupling properties. Phloretin appears to be an active uncoupler; its 6′-glucoside is without effect.     

Isolation and characterization of a novel mung bean protease inhibitor with antipathogenic and anti-proliferative activities

A novel protease inhibitor, designated mungoin, with both antifungal and antibacterial activities, and exhibiting a molecular mass of 10 kDa in SDS-polyacrylamide gel electrophoresis, was isolated from mung bean (Phaseolus mungo) seeds. The isolation procedure involved a combination of extraction, ammonium sulfate precipitation, ion exchange chromatography on CM-Sephadex, and high-performance liquid chromatography (HPLC) on SP-Toyopearl. Its isoelectric point was estimated to be 9.8 by isoelectric focusing. Its N-terminal amino acid sequence was determined to be EMPGKPACLDTDDFCYKP, demonstrating some resemblance to the C-terminal sequences of other protease inhibitors and inhibitor precursors from leguminous plants. It exerted a potent inhibitory action toward a variety of fungal species including Physalospora piricola, Mycosphaerella arachidicola, Botrytis cinerea, Pythium aphanidermatum, Sclerotium rolfsii and Fusarium oxysporum, as well as an antibacterial action against Staphylococcus aureus. In addition, this novel plant protease inhibitor displayed anti-proliferative activity toward tumor cells.     

甘蔗废糖蜜对绿豆插条下胚轴生根的影响

本文研究甘蔗废糖蜜对绿豆插条下胚轴生根的影响,结果表明,1000~7000mg/L浓度范围内的甘蔗废糖蜜能明显增加绿豆插条下胚轴不定根的数目、根长、根干重及生根范围,并促进不定根内可溶性糖含量和不定根系活力提高。