Appearance of a State 1 – State 2 transition during chloroplast development in the wheat leaf: Energetic and structural considerations

The ability of developing chloroplasts to dynamically regulate the distribution of excitation energy between photosystem 1 and photosystem 2, and thus perform a State 1 – State 2 transition, was examined from analyses of chlorophyll fluorescence kinetics in 4- and 8-day-old Triticum aestivum L. cv. Maris Dove leaves grown under a diurnal light regime. Chloroplasts at all stages of development in the two leaf systems could undergo a State 1 – State 2 transition, except those found in the basal 0.5 cm of the 4-day-old leaf. The ability to physiologically modify the excitation energy distribution between the chlorophyll matrices of the two photosystems developed after the development of mature, fully photochemically competent photosystem 2 units and the appearance of excitation energy transfer between photosystem 2 and photosystem 1. Also, changes in the degree of energetic interaction between the two photosystems, in vivo rates of electron transport and the chlorophyll a/b ratio could not be correlated with the appearance of a State 1 – State 2 transition. Ultrastructural studies demonstrated a 32% increase in the degree of thylakoid appression in chloroplasts at the base of the 8-day-old leaf compared to the situation in the basal 0.5 cm of the 4-day-old leaf. This difference in thylakoid stacking can account for the differing abilities of these two tissues to perform a State 1 – State 2 transition when considered in the context of the distribution of the two photosystems within appressed and non-appressed regions of thylakoid membranes.     

Amino acid sequence of a phosphocholine-binding antibody from an immune defective CBA/N mouse employing the T15 VH region associated with unusual DH, JH, and V kappa segments

Mice expressing the xid gene exhibit an altered immune response to phosphocholine (PC)-conjugated keyhole limpet hemocyanin (KLH). Less than 25% of their anti-PC-KLH response is PC specific, and most of these antibodies lack the normally predominant T15 idiotype. These findings suggested that immune defective mice might employ different variable region genes than normal mice in their anti-PC response. To examine this possibility, we characterized by Southern blot analysis the gene family encoding PC-VH regions and determined the amino acid sequence and fine specificity of binding of a T15-, IgG2, PC-specific hybridoma (1B8E5) produced by fusion of the SP2/O cell line and PC-KLH immune CBA/N spleen cells. Southern blot analysis of DNA from CBA/N mice by using a PC-VH probe (S107 VH) revealed a hybridization pattern virtually identical to that of DNA from normal CBA/J mice, indicating that CBA/N mice do not suffer from a gross deletion of PC-VH genes. Analysis of the 1B8E5 antibody reveals that both the binding specificity and relative affinity of this antibody are different from the anti-PC antibodies of the T15, M167-M511, and M603 families. The complete amino acid sequence of the heavy (H) chain variable region shows that 1B8E5 uses a VH segment identical to the allelic form of T15 (C3) but has a unique D region of three amino acids and use the JH1 joining segment. Both the DH and JH regions are unusual when compared to PC-specific antibodies from normal mice, which have a D region composed of five to eight amino acids and use the JH1 joining segment. The amino terminal sequence of the 1B8E5 light (L) chain demonstrates that this anti-PC antibody carries a Vk3 subgroup L chain. Chains from this subgroup have not previously been found in association with PC-binding antibodies. Thus, the Vk, DH, and JH segments expressed in 1B8E5 make this hybridoma unique in terms of the anti-PC antibodies studied to date, and suggests that additional PC-specific antibodies exist in inbred mice that employ "unusual" V gene segments.     

Metabolism of the herbicide bromoxynil by Klebsiella pneumoniae subsp. ozaenae

Enrichment of soil samples for organisms able to utilize the herbicide bromoxynil (3,5-dibromo-4-hydroxybenzonitrile) as a nitrogen source yielded bacterial isolates capable of rapidly metabolizing this compound. One isolate, identified as Klebsiella pneumoniae subsp. ozaenae, could completely convert 0.05% bromoxynil to 3,5-dibromo-4-hydroxybenzoic acid and use the liberated ammonia as a sole nitrogen source. Assays of cell extracts of this organism for the ability to produce ammonia from bromoxynil revealed the presence of a nitrilase (EC 3.5.51) activity. The enzyme could not utilize 3,5-dibromo-4-hydroxybenzamide as a substrate, and no 3,5-dibromo-4-hydroxybenzamide could be detected as a product of bromoxynil transformation. Comparison of related aromatic nitriles as substrates demonstrated that the Klebsiella enzyme is highly specific for bromoxynil.     

Primary productivity of the River Ely,South Wales,U.K.

The present study was carried out on the non-tidal reaches of the River Ely, South Wales, from October 1979 to October 1980. The seasonal variations of the chlorophyll-a content of the phytoplankton was unimodal with one maximum in May and a minimum in December. The chlorophyll-a content varied from 0.277 to 41.259 mg m^?3. The primary productivity showed a bimodal seasonal distribution with two maxima in May and September and lower values throughout the remainder of the year, particularly in winter. The value for the primary productivity varied from 0.269 to 24.302 mg C m^?3 h^?1. A positive correlation was obtained between chlorophyll-a content and primary productivity. The seasonal distribution of the dominant algal species and the saprobity of the River Ely were also studied. The diatom species almost showed a similar seasonal periodicity. Their concentrations were low during the winter months and high during most of the spring and summer months. Many of the dominant diatom species found in the phytoplankton populations were either considered by the other authors as saprobic (Nitzchia palea) or as inhabitants of eutrophic waters (Gomphonema parvulum, Navicula cryptocephala and Synedra ulna). Chlamydomonas spp. were the most abundant green algae followed by Chlorella vulgaris. The effect of sewage effluent disposal and cattle excreta at three of the sampling sites might partly explain the presence of high Chlamydomonas spp. populations at those sites. Physical factors (low flow rates, high transparency and water temperature) and organic pollution at some sampling sites seemed to play an important role in increasing the number of algal species during spring and summer. The non-tidal reaches of the River Ely were found to be β-mesosaprobic above and below Station 5 and α-mesosaprobic at the latter station and therefore, the river can be considered as polluted at Station 5 and mildly polluted at the others.     

Formation of complexes between avidin and β-adrenergic receptors using biotinyl-alprenolol derivatives

The goal of this study was to synthesize biotinylated derivatives of alprenolol, a β-adrenergic antagonist, and to determine whether these ligands could bind simultaneously to both avidin (a biotin-binding protein) and to the β-adrenergic receptor. Such ligands would be useful for β-adrenergic receptor localization and purification, since avidin can be covalently labelled with fluorescent or electron-dense markers or can be linked to solid supports for affinity chromatography. Three biotinyl derivatives of alprenolol were synthesized and characterized. Each derivative bound to avidin and also possesed high affinity for the duck erythrocyte β-adrenergic receptor. Two of the compounds, biotinyl-caproyl-cysteaminyl-alprenolol (BCCA) and biotinyl-dodecanoyl-cysteaminyl-alprenolol (BDCA) had the same affinities for the duck erythrocyte β-adrenergic receptor (membrane-bound or digitonin-solubilized) in the absence and presence of avidin. This indicated that high affinity complexes could be formed between the β-adrenergic receptor and avidin using these bifunctional biotinyl-alprenolol ligands. In contrast, biotinyl-cysteaminyl-alprenolol (BCA), in which the distance between the biotin and alprenolol moieties was shorter, had greatly reduced affinity for the duck erythrocyte β-adrenergic receptor in the presence of avidin. Additional studies showed that BDCA, avidin-BDCA, and ferritin-avidin-BDCA were equally potent in inhibiting the isoproterenol stimulation of cAMP accumulation in intact HeLa cells. The data reported in this paper demonstrate the importance of an appropriate spacer sequence to allow correct apposition of the receptor and avidin molecules, and suggest that BDCA may be a useful probe for β-adrenergic receptor localization and purification.     

The purification and specificity of a neutral endopeptidase from rabbit kidney brush border

1. A neutral peptidase, previously shown to be located in the brush border of the proximal tubule, and assayed by its ability to hydrolyse [(125)I]iodoinsulin B chain was purified from rabbit kidney. 2. The starting material for the purification was a microsomal pellet prepared from a homogenate of cortical tissue. The membrane-bound enzymes were solubilized by treatment with toluene and trypsin. About half the neutral peptidase activity was released by this treatment in a form that no longer sedimented with the microsomal pellet and which penetrated polyacrylamide gels when subjected to disc electrophoresis. Other treatments with detergents or proteolytic enzymes either inactivated the peptidase or failed to convert it into a genuinely soluble form. 3. Chromatography with successive columns of Sephadex G-200, DEAE-cellulose and hydroxyl-apatite yielded an enzyme that was free of other brush-border peptidase activities and which was homogeneous on disc electrophoresis and ultracentrifugation. 4. The purified enzyme attacked [(125)I]iodoglucagon at a rate comparable with that for [(125)I]iodoinsulin B chain. It did not appear to attack proteins (insulin, albumin and casein) that had been similarly iodinated. 5. Unlabelled insulin B chain and unlabelled glucagon were substantially hydrolysed by the endopeptidase, whereas insulin and albumin released only trivial amounts of ninhydrin-reacting material. The resistance of insulin to attack by endopeptidase, even after prolonged incubation, was confirmed by biological and immunoassay. 6. The specificity of the peptidase was determined by analysis of the products after incubating unlabelled insulin B chain, and some oligopeptide substrates, including pentagastrin, with the enzyme. All of the bonds readily cleaved were those involving the alpha-amino group of hydrophobic residues, i.e. x-Leu-, x-Val-, x-Tyr-, x-Phe- and x-Met-, provided that the residues were not C-terminal. 7. The enzyme showed only endopeptidase activity. Substrates suitable for aminopeptidases, carboxypeptidases or esterases were not attacked.     

Effect of Magnesium on Replication of Rhinovirus HGP

It is known that plaque formation by some rhinoviruses is greatly enhanced by increasing the concentration of MgCl(2). The mechanism of this action was studied by investigating the effects of MgCl(2) on rhinovirus HGP adsorption, growth, clumping, thermal stability, and cell susceptibility to viral cytopathic effect. The latent period was at most 7 hr, whether virus was propagated in cells maintained in 0.8 or 30 mm MgCl(2), but virus release was 8- to 310-fold greater in the presence of 30mm MgCl(2), depending on initial multiplicity. Intracellular virus content appeared unaffected by Mg(++) and reached maximal yield (plateau phase) at about 10 hr. Viral adsorption was increased when cells were maintained in 30mm Mg(++). It is likely that the two effects of magnesium, enhanced adsorption and increased virus release, both contribute to enhancement of plaque formation.