The 230-kDa gamete surface protein of Plasmodium falciparum is also a target for transmission-blocking antibodies

Immunization with extracellular sexual stages of the malaria parasites can induce the production of antibodies which block the development of the parasites in the midgut of a mosquito after a blood meal. We have generated a number of monoclonal antibodies against gametes and zygotes of the human malaria Plasmodium falciparum. Two monoclonal antibodies (mAb) reacting with a 230-kDa gamete surface protein (mAb 1B3 and 2B4 both isotype IgG2a) were found to block transmission of P. falciparum to mosquitoes. Blocking was complement dependent and this was verified in vitro by the rapid lysis of newly formed gametes and zygotes in the presence of the mAb and active complement. Both mAb reacted by immunofluorescence with the surface of gametes and zygotes from isolates of P. falciparum from various geographical areas. Each mAb immunoprecipitated a 230-kDa protein from 125I-labeled surface proteins of newly formed gametes and zygotes and immunoblotted a protein doublet of about molecular mass 260 and 230 kDa from gametocytes and gametes of P. falciparum. Only the 230-kDa protein is expressed on the surface of newly formed macrogametes and zygotes. The 230-kDa gamete surface protein forms a molecular complex with two proteins of 48 and 45 kDa. The 48- and 45-kDa gamete surface proteins have previously been shown to be targets of mAb which block infectivity of P. falciparum to mosquitoes. The present study now demonstrates that antibodies against the 230-kDa gamete surface protein block transmission of P. falciparum to mosquitoes. The 230-kDa gamete protein is thus a potential candidate for a gamete vaccine.     

Evidence for negative regulation of T cell growth by low affinity interleukin 2 receptors

Two experimental situations have been studied, and the results provide evidence for a negative regulatory role for the low affinity interleukin 2 receptor (LA-IL 2R). The IL 2-dependent T helper cell line L-14, deprived of IL 2, becomes quiescent and expresses comparable numbers of high affinity IL 2R (HA-IL 2R) and LA-IL 2R. After activation by recombinant IL 2, this cell line preferentially expresses LA-IL 2R. The IL 2 responsiveness of the L-14 cell line was found to vary according to the ratio of LA-IL 2R to HA-IL 2R: the relative predominance of the LA-IL 2R coincides with a hyporeactivity of cells to IL 2. In contrast, a predominance of HA-IL 2R is accompanied by an increase in cellular IL 2 reactivity. Treatment of three IL 2-dependent T cell lines (L-14, HT-2, and C30.1) with limited amounts of recombinant IL 2 and moderate concentrations of anti-IL 2R monoclonal antibodies stimulates T cell growth. This treatment was shown to selectively diminish the expression of membrane LA-IL 2R. The stimulation was attributed to the decrease of expression of LA-IL 2R.     

Relationship between Sodium Influx and Salt Tolerance of Nitrogen-Fixing Cyanobacteria

The relationship between sodium uptake and cyanobacterial salt (NaCl) tolerance has been examined in two filamentous, heterocystous, nitrogen-fixing species of Anabaena. During diazotrophic growth at neutral pH of the growth medium, Anabaena sp. strain L-31, a freshwater strain, showed threefold higher uptake of Na⁺ than Anabaena torulosa, a brackish-water strain, and was considerably less salt tolerant (50% lethal dose of NaCl, 55 mM) than the latter (50% lethal dose of NaCl, 170 mM). Alkaline pH or excess K⁺ (>25 mM) in the medium causes membrane depolarization and inhibits Na⁺ influx in both cyanobacteria (S. K. Apte and J. Thomas, Eur. J. Biochem. 154:395-401, 1986). The presence of nitrate or ammonium in the medium caused inhibition of Na⁺ influx accompanied by membrane depolarization. These experimental manipulations affecting Na⁺ uptake demonstrated a good negative correlation between Na⁺ influx and salt tolerance. All treatments which inhibited Na⁺ influx (such as alkaline pH, K⁺ above 25 mM, NO₃⁻, and NH₄⁺), enhanced salt tolerance of not only the brackish-water but also the freshwater cyanobacterium. The results indicate that curtailment of Na⁺ influx, whether inherent or effected by certain environmental factors (e.g., combined nitrogen, alkaline pH), is a major mechanism of salt tolerance in cyanobacteria.     

Propeptide of human protein C is necessary for gamma-carboxylation

Protein C is one of a family of vitamin K dependent proteins, including blood coagulation factors and bone proteins, that contains gamma-carboxyglutamic acid. Sequence analysis of the cDNAs for these proteins has revealed the presence of a prepro leader sequence that contains a pre sequence or hydrophobic signal sequence and a propeptide containing a number of highly conserved amino acids. The pre region is removed from the growing polypeptide chain by signal peptidase, while the pro region is subsequently removed from the protein prior to secretion. In the present study, deletion mutants have been constructed in the propeptide region of the cDNA for human protein C, and the cDNAs were then expressed in mammalian cell culture. These deletions included the removal of 4, 9, 12, 15, 16, or 17 amino acids comprising the carboxyl end of the leader sequence of 42 amino acids. The mutant proteins were then examined by Western blotting, barium citrate adsorption and precipitation, amino acid sequence analysis, and biological activity and compared with the native protein present in normal plasma. These experiments have shown that protein C is readily synthesized in mammalian cell cultures, processed, and secreted as a two-chain molecule with biological activity. Furthermore, the pre portion or signal sequence in human protein C is 18 amino acids in length, and the pro portion of the leader sequence is 24 amino acids in length. Also, during biosynthesis and secretion, the amino-terminal region of the propeptide (residues from about -12 through -17) is important for gamma-carboxylation of protein C, while the present data and those of others indicate that the carboxyl-terminal portion of the propeptide (residues -1 through -4) is important for the removal of the pro leader sequence by proteolytic processing.     

Phenylglyoxal modification of the Photosystem II reaction center

Time-resolved spectroscopic techniques, including optical flash photolysis and electron spin resonance (ESR), have been used in conjunction with fluorescence-induction and dye-reduction assays to monitor electron transport in Photosystem II (PS II) subchloroplast particles incubated with the covalent modifier, phenylglyoxal. Phenylglyoxal-modified digitonin (D-10) particles from spinach are characterized by a high initial fluorescence yield (Fi) and an abolition of the variable component of fluorescence (Fv); an inhibition of PS-II-mediated reduction of dichlorophenol indophenol (DPIP) by sym-diphenylcarbazide; an abolition of flash-induced absorption transients (t1/2 greater than 2 microseconds) at 820 nm attributed to the primary electron donor, P-680+; the inhibition of photoreduction of the acceptor Qa; and the elimination of the ESR Signal 2s and Signal 2f. These observations suggest the critical participation of specific arginine residues on both the oxidizing and reducing sides of Photosystem II and also implicate phenylglyoxal as a quinone-binding site inhibitor (Golbeck, J.H. and Warden, J.T. (1984) Biochim. Biophys. Acta 767, 263-271).     

Observation on a 65-kilodalton protein isolated from kanagawa positive strains of Vibrio parahaemolyticus

Crude hemolysin from four KP+ strains of Vibrio parahaemolyticus belonging to serotype 02:K3 exhibited a major protein band (molecular weight, 65 kilodaltons (kDa] in addition to a previously known thermostable direct hemolysin band (molecular weight, 21 kDa) in SDS - polyacrylamide slab gel electrophoresis. These strains showed maximum virulence leading to 100% mouse lethality within 2-6 h. It is hypothesized that this 65-kDa protein may play a vital role in the pathogenesis of the disease caused by V. parahaemolyticus.     

Gibberellic acid by solid state fermentation: Consistent and improved yields

Five strains each of Gibberella fujikuroi and Fusarium monoliforme were screened to select G. fujikuroi P-3, a strain capable of giving consistent production of gibberellic acid (GA(3)) by solid state fermentation (SSF). The comparative production of GA(3) by SSF and submerged fermentation (SmF) indicated better productivity with the former technique. The accumulation of GA(3) was 1.626 times higher in the case of SSF. On the basis of available carbohydrates in the media, the percent conversions were 0.096 and 0.156 in SmF and SSF, respectively. The use of coarse wheat bran of the particle size of 0.3-0.4 cm resulted in an increase of 2.5 times in the yield of GA(3). The enrichment of commercial wheat bran with soluble starch gave enhanced accumulation to an extent of 3.5 times. The relation between GA(3) production and cell growth in SSF was similar to that encountered in SmF. The consistent and improved yields to a tune of 1.22 g GA(3) per kilogram dry moldy bran (DMB) establish the potential and feasibility of SSF for the production of GA(3) by G. fujikuroi P-3. On preliminary cost analysis, a net savings of about 60% and 50% on fermentation medium cost and the expenditure on down-stream processing, respectively, as compared to the presently employed SmF technique was evident.     

Putrescine metabolism in excised cotyledons of Pinus radiata cultured in vitro

The metabolism of ¹⁴C-putrescine and the changes in the endogenous concentrations of putrescine, spermidine and spermine were studied when cotyledons of Pinus radiata D. Don were cultured under shoot-forming (SF, + N⁶-benzyladenine) and non-shoot-forming (NSF, - N⁶-benzyladenine) conditions. Differences in the total uptake of ¹⁴C-putrescine during a 2 h pulse feeding were not significant between the SF and NSF cotyledons except on day 3. The maximum uptake of label was on day 3 in the SF cotyledons, which released the highest amount of ¹⁴CO₂ as well. ¹⁴C from the labeled putrescine was incorporated mainly into γ-aminobutyric acid, aspartate and glutamate. High performance liquid chromatography of the endogenous polyamines indicated that spermidine was the most predominant polyamine in the cultured cotyledons of radiata pine. Spermine increased by about 60% in the SF and 25% in the NSF cotyledons between days 0 and 3 of culture.     

Subcellular localization of cyclo-oxygenase-prostaglandin E2 synthetase complex in goat vesicular gland by catalytic activity analysis

The catalytic activity of the cyclo-oxygenase prostaglandin E2 synthetase complex in subcellular organelles of goat vesicular gland was determined. The enzyme activity was found to be located mostly in the rough endoplasmic reticulum and partly in the nuclear membrane; comparatively very little activity could be detected in the smooth endoplasmic reticulum. There was no detectable activity of the enzyme in the plasma membrane.     

Influence of Co2+, Ni2+ and Fe2+ on the production of tetrapyrroles by Methanosarcina barkeri

Summary The selective formation of three tetrapyrroles, Co-containing corrinoids, Ni-containing factor F₄₃₀ and Fe-containing cytochromes (haems) by Methanosarcina barkeri Fusaro (DSM 804) was achieved as a function of the concentrations of Co²⁺, Ni²⁺ and Fe²⁺ in a methanol minimmum medium. It was found that about 70% of the total tetrapyrroles synthesized was excreted into the culture supernatant. Hence, the continuous production of tetrapyrroles in a fixed-bed reactor (supporter: porous diatomaceous clay) was carried out at a dilution rate of 10 day^-1 (850 ml medium/85 ml column/day). The effluent discharged from the reactor contained the excreted tetrapyrroles, the concentrations of which were dependent upon the Co²⁺, Ni²⁺ and Fe²⁺ concentrations in the feed medium. The maximum productivities from the reactor (1 l basis) were 52 M corrinoids/day, 24 M F₄₃₀/day and 8 M haems/day, respectively.