β-Amylolytic activities ofEmericella nidulans (eidam) vuill

Summary Screening of fungal isolates led to selection of a strain ofEmericella nidulans 45 producing exocellular -amylase in a starch medium. Studies of dialysed enzyme and the formulation of the medium for the enzyme production are presented.     

Effects of lanthanum in cellular systems

Lanthanum belongs to the group of elements known as “lanthanons,” which also includes cerium, europium, promethium, and thulium. It is the most electropositive element of the rare earth group, is uniformly trivalent, and is similar in its chemical properties to the alkaline earth elements. The effects of this element and its compounds on cellular systems are of considerable interest because of their increasing use in industry and as a substitute or antagonist for calcium in a variety of cellular reactions. Lanthanum is also being employed extensively in studying anatomical barriers, membrane structure, and subcellular transport systems, particularly the calcium pathway.     

Expression from symbiotic promoters ofRhizobium meliloti inAzotobacter vinelandii andAzospirillum brasilense

InRhizobium meliloti, the promoter P1 of thenif HDK operon, and also the promoter P2, have earlier been shown to be active in the bacteria present in alfalfa root nodules, but not in the bacteria grown aerobically in culture. Here we have looked at the expression from P1 and P2 in two non-symbiotic nitrogen-fixing bacteria,Azotobacter vinelandii andAzospirillum brasilense, using constructions in which the promoters are fused upstream of theβ-galactosidase gene. The promoter P1, but not P2, is active inA. vinelandii, while neither P1 nor P2 is active inAzospirillum brasilense.     

Mini-exon derived RNA gene ofLeishmania donovani: structure,organization and expression

Mini-exon derived RNA is a small nuclear RNA of trypanosomatid protozoa such asLeishmania which donates its 5′-terminal 39 nucleotides to the 5’-ends of cellular messenger RNAs by trans-splicing. We have cloned a mini-exon derived RNA gene fromLeishmania donovani and studied its organization and expression. About 200 copies of the gene per haploid genome are organized as a tandem repeat on a single chromosome. The gene is transcribed as a 95-nucleotide RNA. The first 39 nucleotides of mini-exon derived RNA is also found at the 5′-terminus of a cellular mRNA (Β-tubulin), thus confirming its identity. Sequence analysis of the gene and its flanking regions showed that while classical RNA polymerase II promoter elements such as TATA and CAAT are absent from the 5′-upstream region, intragenic sequence motifs resembling RNA polymerase III promoter elements are present. The implications of this finding for mini-exon derived RNA expression are discussed.     

Oxygen-derived free radicals and hemolysis during open heart surgery

Reperfusion injury occurs during open-heart surgery after prolonged cardioplegic arrest. Cardiopulmonary bypass also is known to cause hemolysis. Since reperfusion of ischemic myocardium is associated with the generation of oxygen free radicals, and since free radicals can attack a protein molecule, it seems reasonable to assume that hemolysis might be the consequence of free radical attack on hemoglobin protein. The results of this study demonstrated that reperfusion following ischemic arrest caused an increase in free hemoglobin and free heme concentrations, simultaneously releasing free iron and generating hydroxyl radicals. In vitro studies using pure hemoglobin indicated that superoxide anion generated by the action of xanthine oxidase on xanthine could release iron from the heme ring and cause deoxygenation of oxyhemoglobin into ferrihemoglobin. This study further demonstrated that before the release of iron from the heme nucleus, oxyhemoglobin underwent deoxygenation to ferrihemoglobin. The released iron can catalyze the Fenton reaction, leading to the formation of cytotoxic hydroxyl radical (OH·). In fact, the formation of OH. in conjunction with hemolysis occurs during cardiac surgery, and when viewed in the light of the in vitro results, it seems likely that oxygen-derived free radicals may cause hemolysis during cardiopulmonary bypass and simultaneously release iron from the heme ring, which can catalyze the formation of OH·.     

Trans replication and high copy numbers of wheat dwarf virus vectors in maize cells.

The replication of shuttle vectors derived from Wheat Dwarf Virus, a monopartite geminivirus, was studied in cultured maize endosperm cells, and in the Black Mexican Sweet (BMS) maize cell line. Using in vivo labeling and DNA methylation analysis, we showed that replication was initiated within 24 hrs after transfection, and did not require cell division in both cell lines. Copy numbers of 30,000 ds DNA molecules per cell were observed in endosperm cells after three days. The replication protein was shown to act in trans, since the wild type gene of the shuttle vector enabled replication-deficient vectors carrying mutated genes to replicate. These properties suggest that WDV may have similar applications in plants as SV40 in mammalian cells.     

Anaerobic growth on elemental sulfur using dissimilar iron reduction by autotrophic Thiobacillus ferrooxidans

Abstract Anaerobic growth on elemental sulfur using dissimilar iron reduction by Thiobacillus ferrooxidans has been demonstrated. The ferric ion reducing activity (FIR) of the anaerobic cells was double that of the aerobic cells. Significant differences in inhibition of FIR by respiratory inhibitors were observed between aerobic and anaerobic cells. A higher amount of cytochrome was detected in anaerobic cells compared to aerobic cells. Absorption minima developed with the addition of ferric sulfate in the dithionite reduced cell suspension demonstrated that the ferric ion could accept electrons from the cytochrome system of this bacterium. The possibility of two different electron transport chains in ferric ion reduction is discussed.     

Study of properties of cholinephosphotransferase from fetal guinea pig lung mitochondria and microsomes

Summary We have reported earlier that cholinephosphotransferase (EC 2.7.8.2) is present in both mitochondria and microsomes of fetal guinea pig lung. This study was designed to compare the properties of mitochondrial and microsomal cholinephosphotransferase in fetal guinea pig lung. Various parameters, such as substrate specificity, K_m values, sensitivity to N-ethylmaleimide, dithiothreitol and trypsin were measured. Both showed significant preference for unsaturated diacylglycerols over saturated diacylglycerols. Data on K_m and V_max indicate that the affinity of this enzyme for different diacylglycerols varies between the two forms. The ID₅₀ values for N-ethylmaleimide were 20 mM and 12.5 mM for the mitochondrial and microsomal form of the enzyme, respectively. Dithiothreitol showed an inhibitory effect on both; however, the mitochondrial form was inhibited less than the microsomal form. The effects of N-ethylmaleimide and dithiothreitol on both forms of enzyme indicated that the microsomal cholinephosphotransferase requires a higher concentration of -SH for its activity than the mitochondrial enzyme does. The enzyme was inhibited by trypsin in both mitochondria and microsome under isotonic condition suggesting that this enzyme is on the outside of the membrane in both endoplasmic reticulum and mitochondria.     

A new allele of the duplicated 27kD zein locus of maize generated by homologous recombination.

The 27kD zein storage protein locus in many inbred lines of maize consists of a tandem duplication of 12kb, with an expressed gene in each repeat, termed A and B. A single-copy allele with only the A gene can be generated from this duplication in particular stocks of the maize inbred line A188 by a mitotic process that includes a crossover at the 3' regions of the two genes (1). Here, we characterize a second single-copy allele with only the B gene, found in different stocks of A188. This allele arises from a homologous recombination at the highly conserved 5' regions of the two repeats, and cloning and sequencing of this allele define the crossover region. The A and B genes in the duplicated allele were previously shown to be expressed at different levels; this difference remains unchanged in either recombinant allele. Therefore, the crossover points of these two recombinant alleles define the borders of cis-acting sequences responsible for the differential expression of the two genes.