Comparison of globulin proteins from Phaseolus vulgaris with those from Vicia faba

Extraction of maturing Phaseolus vulgaris seeds with an ascorbic acid—NaCI medium facilitated the preparation of two globulin fractions which wer     

Lectin variability in Phaseolus coccineus

Lectin variability within Phaseolus coccineus is revealed by non-denatured electrophoretic patterns and immunological labelling of total seed protein extracts, showing that the different cultivars and wild varieties studied can be classified into three main categories according to the number of isolectins (three, two or one) present in each extract. Attempts in the purification of these isolectins were performed on three different affinity systems in which ligands were thyroglobulin (known to purify the P. vulgaris isolectins), pig red cell membrane ghosts (stroma) or antibodies against the P. vulgaris cv. Contender E₂L₂ isolectin. The P. coccineus isolectins exhibit varied affinities towards thyroglobulin and stroma, the cathodic and anodic (pH 4.5) isolectins being respectively retained by the two systems, whereas the antibody affinity system is the only one able to purify the totality of the isolectins present in an extract.     

Protein-polysaccharide linkages in glycoproteins from Phaseolus vulgaris

Serine and hydroxyproline participate in protein-polysaccharide linkages in hydroxyproline-poor glycoproteins from Phaseolus vulgaris cv Pinto. Most substituted hydroxyproline residues contain arabinose, galactose and glucose, but some have arabinose only. Serine residues contain arabinose, galactose and glucose.     

Purification and characterization of the major storage proteins of Phaseolus vulgaris seeds,and their intracellular and cotyledonary distribution

Several extraction and fractionation procedures have been employed to isolate the major storage proteins of mature seeds of Phaseolus vulgaris cv. “Seafarer”; three proteins which were soluble at pH 4,7, and one that was insoluble at that pH were identified. The characteristic subunits of the three pH 4.7 soluble proteins had MW's 50000 and 47000, 32000, and 23000 respectively; those of the pH 4.7 insoluble fractions had MW 60000 and 20000. Amino acid compositions, N-terminal amino acid residues and the presence of carbohydrate in these proteins have been determined. All these proteins occurred in the protein body fraction and their relative amounts were different in the outer and central parts of the cotyledons.     

Isolation and characterization of a lectin from the seeds of Erythrina Edulis

A galactose-specific lectin was isolated from the seeds of Erythrina edulis. The protein was purified by affinity chromatography of the globulin fraction on an allyl-galactoside polyacrylamide gel. The hemagglutination properties, amino acid composition, A₂₈₀, MW of the protein and of its subunits, carbohydrate content, electrophoretic pattern and isoelectric point were determined. Comparison of its properties with those of other Erythrina lectins shows that the protein is a distinct member of this group of lectins.     

Primary metabolites of Phaseolus vulgaris nodules

More ethanol soluble material (carbohydrate and amino nitrogen) was found in both host cell and bacteroid components of Phaseolus vulgaris nodules from plants grown at 28 W/m² than from plants grown at 7 W/m². The range of compounds identified was similar at the two irradiances. On feeding ¹⁴CO₂ to the plant tops at either irradiance the labelling patterns of carbohydrates and organic acids in the nodule host cells and bacteroids suggested that any or all of the following substances could be donated by the host to the bacteroids for general metabolism: sucrose, fructose, glucose, an unidentified carbohydrate, malic acid and an organic acid co-chromatographing with 6-phosphogluconate. Distribution and labelling patterns of nodule amino compounds were consistent with the hypothesis that ammonia is the primary product of nitrogen fixation within bacteroids, and that this ammonia is transported to host cells for assimilation, initially into glutamine and glutamate.     

Early biochemical events during adventitious root initiation in the hypocotyl of Phaseolus vulgaris

Indole butyric acid (IBA) initiates roots in the hypocotyl tissue of Phaseolus vulgaris (French bean). The response is dependent on the concentration of IBA and the duration of exposure to the hormone. IBA enhances the rate of total protein synthesis in ca 30 min after exposure of the hypocotyl segments to the hormone. There is no detectable change in total or poly(A)-containing RNA synthesis in this period although significant increases are seen 2 hr after hormone pre-treatment. The early IBA-mediated increase in protein synthesis (30 min) is not sensitive to Actinomycin D but the antibiotic blocks the increase manifested 2 hr after hormone pre-treatment. Inhibition of early protein synthesis by cycloheximide depresses and delays root initiation. Cytosol prepared from IBA-treated hypocotyl tissue stimulates protein synthesis in vitro to a greater extent than that of the control.     

A cathepsin L-like protease from Strongylusvulgaris: An orthologue of Caenorhabditiselegans CPL-1

Cathespin L-like proteases (CPLs), characterized from a wide range of helminths, are significant in helminth biology. For example, in Caenorhabditiselegans CPL is essential for embryogenesis. Here, we report a cathepsin L-like gene from three species of strongyles that parasitize the horse, and describe the isolation of a cpl gene (Sv-cpl-1) from Strongylusvulgaris, the first such from equine strongyles. It encodes a protein of 354 amino acids with high similarity to other parasitic Strongylida (90-91%), and C.elegans CPL-1 (87%), a member of the same Clade. As S.vulgariscpl-1 rescued the embryonic lethal phenotype of the C.eleganscpl-1 mutant, these genes may be orthologues, sharing the same function in each species. Targeting Sv-CPL-1 might enable novel control strategies by decreasing parasite development and transmission.     

Variation in low molecular weight carbohydrate composition of Phaseolus vulgaris seeds

Variations in the low M_r, carbohydrate composition have been observed in wild forms of common bean (Phaseolus vulgaris) seed. In four of 23 samples, verbascose content in the seeds was quite high and the ratio verbascose—stachyose was more than 1.0. This type of carbohydrate composition was named type A, and has a much higher verbascose content and lower galactinol and stachyose content than the rest of the wild forms, named type B. Although the total and individual carbohydrate content, with the exception of the verbascose content, of the cultivated forms of the common bean were higher than those of wild forms, the carbohydrate composition of the cultivated forms was essentially similar to type B of the wild forms. This carbohydrate composition was considered to be basic to the species.     

Genetic diversity and symbiotic effectiveness of Phaseolus vulgaris-nodulating rhizobia in Kenya

Phaseolus vulgaris (common bean) was introduced to Kenya several centuries ago but the rhizobia that nodulate it in the country remain poorly characterised. To address this gap in knowledge, 178 isolates recovered from the root nodules of P. vulgaris cultivated in Kenya were genotyped stepwise by the analysis of genomic DNA fingerprints, PCR-RFLP and 16S rRNA, atpD, recA and nodC gene sequences. Results indicated that P. vulgaris in Kenya is nodulated by at least six Rhizobium genospecies, with most of the isolates belonging to Rhizobium phaseoli and a possibly novel Rhizobium species. Infrequently, isolates belonged to Rhizobium paranaense, Rhizobium leucaenae, Rhizobium sophoriradicis and Rhizobium aegyptiacum. Despite considerable core-gene heterogeneity among the isolates, only four nodC gene alleles were observed indicating conservation within this gene. Testing of the capacity of the isolates to fix nitrogen (N₂) in symbiosis with P. vulgaris revealed wide variations in effectiveness, with ten isolates comparable to Rhizobium tropici CIAT 899, a commercial inoculant strain for P. vulgaris. In addition to unveiling effective native rhizobial strains with potential as inoculants in Kenya, this study demonstrated that Kenyan soils harbour diverse P. vulgaris-nodulating rhizobia, some of which formed phylogenetic clusters distinct from known lineages. The native rhizobia differed by site, suggesting that field inoculation of P. vulgaris may need to be locally optimised.     

Cyclic nucleotide phosphodiesterase activity in Phaseolus vulgaris

Centrifugal fractionation showed that 70% of the cyclic nucleotide phosphodiesterase activity of Phaseolus vulgaris seedlings is recovered in the 1     

Enzymic formation of carbonyls from linoleic acid in leaves of Phaseolus vulgaris

Homogenization of Phaseolus vulgaris leaves at acid pH results in the evolution of hexanal, cis-3- and trans-2-hexenal. With cell-free extracts of leaves, linoleic and linolenic acids are enzymically converted to their hydroperoxides (predominantly the 13-hydroperoxide isomers) and to hexanal or hexenal respectively. Activity was highest in young, dark-green leaves and was stimulated by Triton X-100. Oleic acid is not a substrate for these reactions. Both 9- and 13-hydroperoxides were cleaved to carbonyl fragments and are proposed as intermediates in the formation of volatile aldehydes and non-volatile ω-oxoacids in P. vulgaris leaves. Properties of the enzyme systems are described.     

The role of copper and quaternary structure on the conformational properties of Octopus vulgaris hemocyanin

Some structural properties of Octopus vulgaris hemocyanin have been investigated by fluorescence spectroscopy. The three-dimensional structure of Octopus hemocyanin is remarkably tight, resulting in a deep burial of almost all the tryptophyl residues of the protein. The hemocyanin conformation has been studied in the two main aggregation states (11 S, 50 S) of the protein, and with respect to the presence or absence of copper in the active site. Upon changing the pH of the solution, Octopus hemocyanin in the 50 S aggregation state can assume at least three different conformations. During the transition between each conformation the fluorescence quantum yield changes, but the environment of tryptophans does not change. Dissociation of the protein from 50 S to 11 S strongly enhances its susceptibility toward denaturating agents such as pH or temperature, and modifies the effects of fluorescence quenchers such as acrylamide. Moreover, these effects are more pronounced when copper is removed from the active site. A comparative analysis of the results shows that the subunit-subunit interactions exerted within the 50 S species are more important in the maintenance of the conformational stability than the copper ions present in the active sites. This behavior can be accounted for by the large amount of Ca(II) ions linked to 50 S hemocyanin.     

Expression and interaction of the CBLs and CIPKs from immature seeds of kidney bean (Phaseolus vulgaris L.)

Protein phosphorylation plays a key regulatory role in a variety of cellular processes. To better understand the function of protein phosphorylation in seed maturation, a PCR-based cloning method was employed and five cDNA clones (pvcipk1-5) for protein kinases were isolated from a cDNA library prepared from immature seeds of kidney bean (Phaseolus vulgaris L.). The deduced amino acid sequences showed that the five protein kinases (PvCIPK1-5) are members of the sucrose non-fermenting 1-related protein kinase type 3 (SnRK3) family, which interacts with calcineurin B-like proteins (CBLs). Two cDNA clones (pvcbl1 and 2) for CBLs were further isolated from the cDNA library. The predicted primary sequences of the proteins (PvCBL1 and 2) displayed significant identity (more than 90%) with those of other plant CBLs. Semi-quantitative RT-PCR analysis showed that the isolated genes, except pvcbl1, are expressed in leaves and early maturing seeds, whereas pvcbl1 is constitutively expressed during seed development. Yeast two-hybrid assay indicated that among the five PvCIPKs, only PvCIPK1 interacts with both PvCBL1 and PvCBL2. These results suggest that calcium-dependent protein phosphorylation-signaling via CBL-CIPK complexes occurs during seed development.     

The amino acid sequence of cytochrome c from niger-seed, Guizotia abyssinica

The amino acid sequence of cytochrome c from niger-seed has been determined by sequence analysis of chymotryptic and tryptic peptides using the dansyl-phenylisothiocyanate method and by qualitative analysis of peptide composition by the dansyl method. Although the spectral ratios indicated the protein was not completely pure, no indication of impurity was found during the sequence analysis and no peptides in addition to those given here were obtained. In certain cases the alignment of peptides was by homology with other cytochromes c. Four residues in the proposed sequence, alanine-1, cysteine-25, histidine-26 and lysine-61 were identified only from peptide compositions. The amino-terminus of the protein is acetylated. The sequence contains two residues of ϵ-N-trimethyllysine.     

Biosynthesis of the phytoalexin phaseollin in Phaseolus vulgaris

Feeding experiments in CuCl₂-treated French bean (Phaseolus vulgaris) seedlings have demonstrated that labelled 2′,4′,4-trihydroxychalcone, daidzein, 7,2′,4′-trihydroxyisoflavone, 3,9-dihydroxypterocarpan and phaseollidin are all good precursors of the pterocarpan phytoalexin phaseollin. These compounds represent a logical sequence in the biosynthetic pathway to phaseollin.     

Occurrence of guanosine 3′,5′-cyclic monophosphate (Cyclic GMP) and associated enzyme systems in Phaseolus vulgaris

Cyclic GMP, isolated from Phaseolus vulgaris, has been unequivocally identified by NMR and FAB-mass spectrometry with MIKES-scanning. Radioimmunoas     

Epoxidase/epoxide hydrase activity in cell cultures of Phaseolus vulgaris

The presence of epoxidase and epoxide hydrase enzymes in cell suspension culture of Phaseolus vulgaris is demonstrated. Results indicate high levels of enzyme activity using stilbene and stilbene oxide as substrates.     

Starch synthesis in isolated Phaseolus vulgaris chloroplasts

Isolated bean (Phaseolus vulgaris) chloroplasts were used to investigate the mode of synthesis of transitory amylose and amylopectin from ADP-glucose. Pulse chase experiments showed that labelled glucose in amylose decreased when chased with cold substrate as compared to controls. A significant portion of this decrease appeared in the amylopectin fraction indicating that amylopectin was formed from amylose. However, time course experiments showed that the rate of amytopectin synthesis is higher than that of amylose at the early stages of incubation, suggesting a certain degree of independent synthesis of the two fractions. High concentration of citrate increased the rate of amylopectin synthesis.     

Metabolism of [6-14C]orotate by shoots of Pisum sativum, Phaseolus vulgaris and Lathyrus tingitanus

Incorporation of radioactivity from [6-¹⁴C]orotate into the pyrimidine constituents of shoots of Pisum sativum, Phaseolus vulgaris and Lathyrus tingitanus was examined with special reference to the unusual pyrimidine constituents. With each species, although 80% of the orotate supplied was catabolized to β-alanine, all the pyrimidine derivatives became radioactively labelled. With Pisum, the major part of the radioactivity incorporated into pyrimidines was located in UMP and the uracil derivatives, including the uracilyl amino acids willardiine and isowillardiine. With Phaseolus, UMP and the uracil derivatives were again the major radioactive products; incorporation of radioactivity into 5-ribosyluracil (pseudouridine), which accumulates in Phaseolus tissues, was comparable to the incorporation into orotidine and twice that found in cytidine. Lathyrus incorporated a substantially larger part of the presented [6-¹⁴C] orotate into pyrimidine derivatives than did the other two species. CMP was the most highly radioactive product, followed next by lathyrine and UMP. Surprisingly, 20% of the total radioactivity incorporated into pyrimidines by Lathyrus was located in the pyrimidine amino acid lathyrine. This confirms previous evidence that lathyrine is essentially a product of the orotate pathway. The overall recovery of radioactivity in all three species was 93–95%. The data emphasize the necessity of including the less common pyrimidine constituents, as well as the common ones, in quantitative studies of pyrimidine metabolism in plants.