Endogenous evolution of HCN during pre-germination periods in many seed species

Both cyanogenic ( Malus pumila Mill) and acyanogenic ( Oryza sativa L., Hordeum vulgare L., Zea mays L., Glycine max Merr., Lactuca sativa L., and Xanthium pennsylvanicum Wallr. etc.) seeds evolve HCN gas during the early periods of water imbibition. All tested seeds contained reserve cyanogens which liberated HCN upon hydrolysis with H₂SO₄ and with β-glucosidase and/or lipase. The amounts of liberated HCN were roughly comparable to those of unidentified cyanogens. It is thus conceivable that the cyanogens within seeds are available as precursors for free HCN evolved in the pre-germination period. The amounts of HCN evolved in the acyanogenic seeds were only 0.002 to 1% of that in apple, but the contents of the cyanogenic compounds in rice and cocklebur increased temporarily during the pre-germination period, then decreased and, finally, disappeared completely with the start of germination.     

Presence of β-cyanoalanine synthase in unimbibed dry seeds and its activation by ethylene during pre-germination

Evolution of HCN from both rice ( Oryza sativa ) and cocklebur ( Xanthium pennsylvanicum ) seeds increased during a pre-germination period and preceded the evolution of (C₂H₄). These two species were adopted as the representatives of starchy and fatty seeds, respectively. Ethylene promotes seed germination of many species. However, HCN evolution declined abruptly when the radicles emerged and before the peak in C₂H₄ evolution. More-over, both rice and soybean ( Glycine max ) seeds showed some activity of β-cyanoalanine synthase (CAS, EC 4.4.1.9) even in the unimbibed dry state. The activities of CAS in the lower seed of cocklebur and in soybean seeds increased rapidly after emergence of the radicle. However, the CAS of rice seeds, with high activity in the dry state, exhibited a bimodal change, gradually decreasing until radicle emergence had occurred, but then increaing. It is thus likly that HCN evolution during initial imbibition may be derived from cyanogenic reserves and controlled by both pre-existing and subsequently-developing CAS. The exogenous application of C₂H₄ stimulated the activities of CAS in both rice and upper cocklebur seeds and reduced their cyanogen contents. Therefore, the decline of HCN evolution after germination seems to be due to the increased activities of CAS by endogenously produced C₂H₄.     

The presence of two types of β-cyanoalanine synthase in germinating seeds and their responses to ethylene

Germinating seeds of many species contain two types of β-cyanoalanine synthase (CAS, EC 4.4.1.9) that convert HCN to β-cyanoalanine. One is cytoplasmic CAS (cyt-CAS), which is precipitated by 50 to 60% (NH₄)₂SO₄ and has a pH optimum of 10.5. Cytoplasmic CAS is present at high levels in dry seed and its activity does not increase during imbibition. The activity of cyt-CAS is not affected by exogenously applied ethylene (C₂H₄), except in rice ( Oryza sativa cv. Sasanishiki). The second type of CAS found in seed is mitochondrial CAS (mit-CAS), which is precipitated by 60 to 70% (NH₄)₂SO₄ and has a pH optimum of 9.5. Mitochondrial CAS is present at low levels in dry seed, and its activity increases greatly during imbibition in the seeds of all species tested. Exposure to C₂H₄ stimulated mit-CAS activity in seeds of rice, barley ( Hordeum vulgare cv. Hadakamugi). cucumber ( Cucumis sativus cv. Kagafushinari) and cocklebur ( Xanthium pennsylvanicum ). The increase in the mit-CAS activity in cocklebur in response to C₂H₄ commenced alter a lag period of 2 to 3 h when the duration of soaking was short (16 h), but commenced without a lag period when the seeds were soaked for three months. Application of both chloramphenicol and cycloheximide to the axial and cotyledonary tissues of cocklebur seeds strongly inhibited growth as well as the increase in mit-CAS activity. It is postulated that the mit-CAS is synthesized de novo during imbibition and that its activity is regulated by C₂H₄, CO₂ which also promotes seed germination in some species, was ineffective m stimulating mit-CAS activity in cocklebur seeds.     

A bimodal germination response to temperature in cocklebur seeds. I. Cyanide-sensitive and cyanide-resistant respirations

Abstract. Pre-imbibed cocklebur ( Xanthium penn-sylvanicum Wallr.) seeds displayed bimodal germination-temperature responses with two optima at 8 and 33° C. Such germination responses occurred subsequent to bimodal respiration-temperature upsurges at lower and higher temperature regions. At lower temperatures, cocklebur seeds respired predominantly through a cyanide-sensitive cytochrome pathway. A rise in temperature resulted in a marked increase in flux via an alternative pathway, a propyl gallate- (PG) or benzohydroxamic-acid- (BHAM) sensitive pathway, thus resulting in an increase in the ratio of this pathway relative to the cytochrome pathway. Both an increased capacity for the alternative pathway and an increase in the ratio of this pathway to the cytochrome pathway were obtained when pre-imbibed seeds were exposed to either 8 or 33°C for a short period. The effects of low temperature were reduced as the exposure time was prolonged beyond 3d, resulting in a reduction in germination. Neither PG nor BHAM had an inhibitory effect on the chilling-induced germination, but the germination-stimulating effect of high temperatures was less pronounced in the presence of PG or BHAM. At high temperatures, on the other hand, KCN and NaN₃ were ineffective or, rather, slightly inhibited germination. It was thus concluded that low and high temperatures exert their germination-stimulating effects by an essentially similar manner which increases fluxes both via the cytochrome pathway and, especially, via the alternative pathway and, as a result, raises the ratio of the latter to the former.     

A bimodal germination response to temperature in cocklebur seeds. II. ATP and adenylate pool

Abstract. The mechanism involved in a bimodal germination-temperature response in pre-soaked cocklebur (Xanthium pennsylvanicum Wallr.) seeds was studied with special reference to adenylate metabolism. Exposure to either low (optimal at 8°C) or high (optimal at 34°C) temperature which was effective in inducing the germination of the seeds brought about the accumulation of ATP in them. The ATP level remained unchanged at temperatures around 23°C. Pretreatment with KCN, stimulating germination even at 23°C, subsequently increased the ATP content, total adenylate pool and energy charge (EC) in the axial tissue prior to germination above those of the untreated controls. The lower the treatment temperature, the greater the inhibitory effect of KCN on ATP formation. An increase in germination following an increasing duration of pre-soaking at 8°C was comparable to increasing both the ATP content and total adenylate pool of axes, but not the EC value. Similarly, changes in germination following an increased exposure duration at 8°C correlated with changes in ATP content rather than EC value in the axes. Unlike the case of chilling, an increase in ATP level in response to 34°C was greater in the early period of water imbibition, during which times its germination-stimulating effect appeared more striking than in the later period, and it occurred without a concomitant rise in EC value because of the increased supply of AMP. Such a supply of AMP was reduced in the presence of benzohydroxamic acid or propyl gallale, inhibitors of an alternative respiratory pathway. It was thus concluded that both low temperature, coupled with warm temperature, and high temperature, by itself, can induce seed germination by increasing the ATP level as well as the total adenylate pool, but not the EC value, in the axial tissue. Further, that increases in both the ATP level and the adenylate pool especially are required for seed germination to proceed, probably depending on the activities of the cytochrome and alternative respiration pathways, respectively.     

α-Aminoisabutyric acid, propyl gallate and cobalt ion and the mode of inhibition of ethylene production by cotyledonary segments of cocklebur seeds

Using cotyledonary segments of cocklebur ( Xanthium pennsylvanicum Wallr. ) seeds, the inhibitory effect of α-aminoisobutyric acid (AIB) on ethylene production was compared with that of propyl gallate and CoCl₂. Of these inhibitors only AIB was effective in causing the accumulation of endogenous free 1-aminocyclopropane-l-carboxylic acid (ACC) in the tissue. The degree of inhibition of ethylene production by AIB decreased markedly with increasing concentrations of pre-loaded ACC, while the inhibition by propyl gallate and CoCl₂ changed little. Kinetic analysis showed that AIB competitively inhibited the conversion of pre-loaded ACC to ethylene, but propyl gallate and CoC_l2 did not. Short-chain organic acids and analogues of AIB, such as acetic, propionic, butyric and cyclopropanecarboxylic acids, did not inhibit ethylene production by the segments. Thus, additional support for the competitive mode of inhibitory action of AIB on the conversion of free ACC to ethylene was provided.
A conjugated hydrolysable ACC was found to be present in abundance in cotyledons of this seed. However, its content in the tissue was hardly affected by treatment with the three inhibitors and by administration of exogenous ACC, suggesting that the conjugated ACC was not directly involved in ethylene production.     

β-Glucosidase from the cellulolytic system of Alternaria alternata autolyzed cultures

Abstract A β-glucosidase from centrifugated autolyzed cultures of Alternaria alternata has been purified 71 times by Sephadex G-200, CM-Biogel A and DEAE-Biogel A successively. The enzyme is a glycoprotein with 16% sugar and a M _r of 160 000, formed by two subunits of 60 000 and 80 000. The enzyme has optimum pH of 5 units and optimum reaction temperature of 50°C, being stable in a pH range of 3–8 and 0 to 60°C. The enzyme hydrolyzes different substrates showing maximum affinity and maximum hydrolysis velocity on cellobiose. The β-glucosidase is inhibited by gluconolactone but not by 10 mM glucose.     

Reversal of ethylene action on cocklebur seed germination in relation to duration of pre-treatment soaking and temperature

Abstract At 23°C, both C₂H₄ and CO₂ stimulated the germination of freshly imbibed upper cocklebur (Xanthium pennsylvanicum Wallr.) seeds, but C₂H₄, unlike CO₂, changed to an inhibitor of germination under some soaking conditions. However, when seeds were pre-soaked for more than several hours at 23 °C prior to treatment, C₂H₄ strongly inhibited their germination at 33 °C, the degree of inhibition increasing with the duration of pre-soaking. Maximum inhibition occurred at 1–3 cm³ m^?3 C₂H₄ when seeds were pre-soaked for 1 week; further increases of C₂H₄ concentration and pre-soaking period decreased the inhibitory effect. C₂H₄ was synergistic with CO₂ when C₂H₄ promoted germination, whereas it was antagonistic when inhibitory. Such a transition of the C₂H₄ action occurred at ca. 27 °C. Also 1-andnocyclopropane-1-carboxylic acid, a C₂H₄ precursor, inhibited the germination of pre-soaked seeds at 33 °C, although it promoted the germination at 23 °C. When pre-soaked seeds were prepared for germination by chilling at 8 °C for 3 d, the inhibitory effect of C₂H₄ on the subsequent germination was manifested even at 23 °C. The reversal of the C₂H₄ action from promotion to inhibition in cocklebur seed germination is discussed in relation to the engagement of two respiratory pathways in the imbibed seeds.     

Cyanogenic glycosides and their metabolic enzymes in barley, in relation to nitrogen levels

The possible role for cyanogenic glycosides as nitrogen storage compounds was studied in barley, Hordeum vulgare (cv. Golf), cultivated under different nitrogen regimes. Cyanogenic glycosides were absent in seeds and roots but were synthesized in seedlings where they accumulated at a level of about 150 nmol shoot⁻¹ in control plants and 110 nmol shoot⁻¹ in nitrogen-starved plants. An enzyme involved in the breakdown of cyanogenic glycosides, β-glucosidase (EC 3.2.1.-) exhibited high activity in seeds and was also detected in roots and shoots. The activity of β-cyanoalanine synthase (EC 4.4.1.9), which is involved in the metabolism of HCN, was low in seeds but very high in roots and shoots. There was no correlation between the activities of the two enzymes and the content of cyanogenic glycosides or nitrogen. The relative content of nitrogen in cyanogenic glycosides never exceeded 0.3% of total nitrogen, and the amount of cyanogenic glycosides decreased at a low rate even at a stage when nitrogen limitation inhibited growth.     

β-Glucosidase and β-Galactosidase in Primary Cultures of Rat Astrocytes: Comparison to the Brain Enzymes

In primary astrocyte cultures beta-glucosidase (EC 3.2.1.21) and beta-galactosidase (EC 3.2.1.23) showed pH optima and Km values identical to rat brain enzymes, using methylumbelliferyl glycosides and labeled gluco- and galactocerebrosides as substrates. The activities of both glycosidases increased in culture up to 3-4 weeks. In rat brain only galactosidase increased; glucosidase activity declined between 12-20 days after birth. The specific activities were two- to sixfold higher in astrocyte cultures than in rat brain. These activities were not due to uptake of enzymes from the growth medium. Secretion of beta-galactosidase, but not beta-glucosidase nor acid phosphatase could be demonstrated. These results support the suggestion of a degradative function for astrocytes in the brain.     

Expression of β-galactosidase multiple forms during barley (Hordeum vulgare) seed germination. Separation and characterization of enzyme isoforms

β-Galactosidase (EC 3.2.1.23) activity in barley ( Hordeum vulgare ) seedlings increases moderately during the first stages of germination. The level of activity in the whole seedling is the result of increasing activity of β-galactosidase in the roots and shoots and of declining enzyme activity in the grain. β-Galactosidase was purified during different developmental stages and from various parts of the barley seedling using affinity chromatography and was resolved into multiple forms by isoelectric focusing on polyacrylamide gels. The expression of the isoforms was shown to be under temporal and tissue-specific control. Four sets of isozymes were separated by DEAE-cellulose chromatography and were shown to be functionally similar. β-Galactosidase isoforms also exhibit size microheterogeneity, the more acidic entities having higher molecular masses. The differences in molecular weight are mainly restricted to the size of the small subunit. Multiplicity can not be attributed to glycosylation, since treatment of the enzyme preparation with N- or O-glycanase did not alter the isoelectric points or the molecular weights of the isoforms.     

Localisation of β-oxidation enzymes in peroxisomes of rice coleoptiles

Enzymes of the β-oxidation pathway in rice ( Oryza sativa L., cv. Arborio) coleoptiles were investigated. The coleoptiles contain acyl-CoA oxidase (EC 1.3.99.3), 3-hydroxyacyl-CoA dehydrogenase (EC 1.1.1.35), enoyl-CoA hydratase (EC 4.2.1.17) and thiolase (EC 2.3.1.9). Analysis of coleoptile homogenates by sucrose density fractionation showed a preferential distribution of these enzymes in the unspecialized peroxisomes. The enzymatic activity found in the mitochondrial fraction was due to peroxisomal contamination since electron micrographs show the peroxisomes to be intact and pure whereas the mitochondrial fraction was contaminated by other organelles. It appears that the β-oxidation pathway is localized in the unspecialized peroxisomes of rice coleoptiles, extending the number of plant species in which such a localization has been observed.     

Immunohistochemical Localization of G Protein β1, β2, β3, β4, β5, and γ3 Subunits in the Adult Rat Brain

Abstract: The regional distributions of the G protein β subunits (Gβ1–β5) and of the Gγ3 subunit were examined by immunohistochemical methods in the adult rat brain. In general, the Gβ and Gγ3 subunits were widely distributed throughout the brain, with most regions containing several Gβ subunits within their neuronal networks. The olfactory bulb, neocortex, hippocampus, striatum, thalamus, cerebellum, and brainstem exhibited light to intense Gβ immunostaining. Negative immunostaining was observed in cortical layer I for Gβ1 and layer IV for Gβ4. The hippocampal dentate granular and CA1–CA3 pyramidal cells displayed little or no positive immunostaining for Gβ2 or Gβ4. No anti-Gβ4 immunostaining was observed in the pars compacta of the substantia nigra or in the cerebellar granule cell layer and Purkinje cells. Immunoreactivity for Gβ1 was absent from the cerebellar molecular layer, and Gβ2 was not detected in the Purkinje cells. No positive Gγ3 immunoreactivity was observed in the lateral habenula, lateral septal nucleus, or Purkinje cells. Double-fluorescence immunostaining with anti-Gγ3 antibody and individual anti-Gβ1–β5 antibodies displayed regional selectivity with Gβ1 (cortical layers V–VI) and Gβ2 (cortical layer I). In conclusion, despite the widespread overlapping distributions of Gβ1–β5 with Gγ3, specific dimeric associations in situ were observed within discrete brain regions.     

Purification of a β-galactosidase from rice shoots and its involvement in hydrolysis of the natural substrate in cell walls

Several glycosidase and glycanase activities have been detected in homogenates of rice ( Oryza sativa L. cv. Nipponbare) shoots after successive extraction with K-phosphate (pH 7. 0) and buffer containing 3 M LiCl. The major β-D-galactosidase (EC 3. 2. 1. 23) present in the buffer-soluble protein fraction was purified to electrophoretic homogeneity by a combination of chromatographic techniques including DEAE-Sepharose CL-6B, Sephacryl S-200HR and p -aminophcnyl-β-D-thiogalactopyranoside–Sepharose 4B. Analysis by denaturing gel electrophoresis revealed a single polypeptide chain with an apparent molecular mass of 42 kDa. Similar to the value of 40 kDa estimated for the native protein by gel-permeation. The isoelectric point was pH 6. 0. The K_m and V_max values for p -nitrophenyl (PNP)-β-D-galactopyra-noside were 0. 63 m M and 0. 32 mmol (mg protein)⁻¹ h⁻¹, respectively. Maximum activity in McIlvaine buffer occurred at pH 3. 4, and the activity was inhibited by Ag²⁺, Cu²⁺. Hg²⁺, p -chloromercuribenzoate (PCMB) and D-galactono-l,4-lactone. The enzyme hydrolyzed larchwood arabinogalactan in an exo-fashion, and acted weakly on arabinosyl and galactosyl residue-rich polymer of pectic polysaccharides and cell walls from rice shoots.     

Effect of GM-CSF on cytokine induction by soluble β-glucan SCG in vitro in β-glucan-treated mice

SCG is a 6-branched 1,3-β- d -glucan, which are major cell wall structural components in fungi. Leukocytes from DBA/1 and DBA/2 mice are highly sensitive to SCG, producing cytokines such as GM-CSF, IFN-γ, TNF-α and IL-12p70, but not IL-6. GM-CSF plays a key biological role in this activity. In the present study, we examined the effect of giving i.p. SCG to DBA/2 mice on cytokine production in vitro . SCG was given i.p. to DBA/2 mice on day 0. Splenocytes were prepared on day 7 and cultured in the presence of SCG in vitro . The levels of cytokine production induced by SCG in vitro were lower in the cells from SCG-treated mice than in control mice. Expression of the β-glucan receptor, dectin-1, in SCG-treated mice was comparable with that shown in control mice. However, the consumption of exogenously added rmGM-CSF in vitro was observed in SCG-treated mice. The addition of a large amount of rmGM-CSF to the culture medium resulted in larger amounts of TNF-α and IL-6 in SCG-treated mice than in normal mice. These results suggested that GM-CSF was closely related with the reactivity of β-glucan. Giving SCG increased the number of macrophages and granulocytes in the spleen. These results suggested that in SCG-treated mice, a change of cell population would be related to modulation of the profile of cytokine production induced by SCG in vitro .     

Inhibition of Tyrosine Aminotransferase by β-N-Oxalyl-l-α,β-Diaminopropionic Acid, the Lathyrus sativus Neurotoxin

Abstract: Species differences in susceptibility are a unique feature associated with the neurotoxicity of β-N-oxalyl-l -α,β-diaminopropionic acid (l -ODAP), the Lathyrus sativus neurotoxin, and the excitotoxic mechanism proposed for its mechanism of toxicity does not account for this feature. The present study examines whether neurotoxicity of l -ODAP is the result of an interference in the metabolism of any amino acid and if it could form the basis to explain the species differences in susceptibility. Thus, Wistar rats and BALB/c (white) mice, which are normally resistant to l -ODAP, became susceptible to it following pretreatment with tyrosine (or phenylalanine), exhibiting typical neurotoxic symptoms. C57BL/6J (black) mice were, however, normally susceptible to l -ODAP without any pretreatment with tyrosine. Among the various enzymes associated with tyrosine metabolism examined, the activity of only tyrosine aminotransferase (TAT) was inhibited specifically by l -ODAP. The inhibition was noncompetitive with respect to tyrosine (K_i = 2.0 ± 0.1 mM) and uncompetitive with respect to α-ketoglutarate (K_i = 8.4 ± 1.5 mM). The inhibition of TAT was also reflected in a marked decrease in the rate of oxidation of tyrosine by liver slices, an increase in tyrosine levels of liver, and also a twofold increase in the dopa and dopamine contents of brain in l -ODAP-injected black mice. The dopa and dopamine contents in the brain of only l -ODAP-injected white mice did not show any change, whereas levels of these compounds were much higher in tyrosine-pretreated animals. Also, the radioactivity associated with tyrosine, dopa, and dopamine arising from [¹⁴C]tyrosine was twofold higher in both liver and brain of l -ODAP-treated black mice. Thus, a transient increase in tyrosine levels following the inhibition of hepatic TAT by l -ODAP and its increased availability for the enhanced synthesis of dopa and dopamine and other likely metabolites (toxic?) resulting therefrom could be the mechanism of neurotoxicity and may even underlie the species differences in susceptibility to this neurotoxin.     

Thiamine increases β-glucosidase production in the newly isolated strain of Fomitopsis pinicola

Aims:  To isolate a high β-glucosidase (BGL)-producing strain and to optimize BGL production in the isolated strain.
Methods and Results:  A high BGL-producing strain was isolated and identified as Fomitopsis pinicola KMJ812 based on its morphology and a comparison of sequence of its internal transcribed spacer rDNA gene. To increase BGL production, F. pinicola was supplemented with various vitamins. Supplementation with thiamine (20 mg l⁻¹) improved BGL production in F. pinicola cultures by 3·7-fold to give a specific activity of 114·4 μmol min⁻¹ mg⁻¹ protein, one of the highest among BGL-producing micro-organisms. The increased production of BGL in the thiamine-supplemented culture was confirmed by 2D electrophoresis followed by MS/MS sequencing. The BGL purified from F. pinicola culture showed the highest catalytic efficiency ever reported.
Conclusion:  Supplemental thiamine remarkably increased BGL production by a novel BGL-producing strain, F. pinicola KMJ812.
Significance and Impact of the Study:  Our results provide a high BGL-producing strain and the production media for BGL production, and should contribute to better industrial production of glucose via biological processes.     

Early Effects of β,β'-Iminodipropionitrile on Tubulin Solubility and Neurofilament Phosphorylation in the Axon

Abstract: To elucidate the role of neurofilaments in microtubule stabilization in the axon, we studied the effects of β,β'-iminodipropionitrile (IDPN) on the solubility and transport of tubulin as well as neurofilament phosphorylation in the motor fibers of the rat sciatic nerve. IDPN is known to impair the axonal transport of neurofilaments, causing accumulation of neurofilaments in the proximal axon and segregation of neurofilaments to the peripheral axoplasm throughout the nerve. Administration of IDPN at various intervals after radioactive labeling of the spinal cord with l -[³⁵S]methionine revealed that transport inhibition occurred all along the nerve within 1–2 days. Transport of cold-insoluble tubulin, which accounts for 50% of axonal tubulin, was also affected. A significant increase in the proportion of cold-soluble tubulin was observed, reaching a maximum at 3 days after IDPN treatment and returning to the control level in the following weeks. Preceding this change in tubulin solubility, a transient decrease in the phosphorylation level of the 200-kDa neurofilament protein was detected in the ventral root using phosphorylation-dependent antibodies. These early changes agreed in timing with the onset of segregation and transport inhibition, suggesting that interaction between neurofilaments and microtubules possibly regulated by phosphorylation plays a significant role in microtubule stabilization.     

Role of β-amylase in starch metabolism during soybean seed development and germination

Differences in starch metabolism during seed development and germination of two soybean [ Glycine max (L.) Merrill] genotypes with normal seed β-amylase activity ['Williams' ( Sp 1^b and 'Altona' ( Sp 1^b)] and two soybean genotypes with undetectable seed β-amylase activity ['Chestnut' ( Sp 1^au) and 'Altona' ( Sp 1)] were investigated. Starch and soluble sugar profiles were essentially the same during seed development and germination. Total amylase activity of Williams and Altona ( Sp 1^b) peaked just prior to seed maturity and then dropped off slowly; whereas, the total amylase activity of Chestnut and Altona ( sp 1) was very low throughout seed development and germination. The differences in amylase activity between Altona ( Sp 1 ^b) and Altona ( sp 1) was also seen in leaves. α-Amylase activity was similar in the four genotypes when β-amylase was inhibited with Hg²⁺ but was higher in the two genotypes with normal β-amylase activity when β-amylase was inhibited with heat plus Ca²⁺. Low levels of starch phosphorylase activity were detected throughout seed development and germination, and the activity was similar in three of the genotypes and higher in Altona ( sp 1).
The protein, oil and oligosaccharide contents of mature seeds of the four genotypes were similar. Altona ( sp 1 ^b) and ( sp 1), which appear to be near isogenic lines, were not different in any morphological character or yield.
Altona ( Sp 1 ^b) showed greater hydrolysis of soybean seed starch than Altona ( sp 1), but the evidence indicates that the mutation resulting in greatly reduced or missing β-amylase activity has no effect on starch metabolism of developing and germinating soybean seeds.