Construction and expression of synthetic DNA fragments coding for polypeptides with elevated levels of essential amino acids

Summary Polypeptides, with elevated levels of essential amino acids, could be useful as partial protein supplements to food and feeds. To obtain DNA fragments coding for these polymers, oligonucleotides were constructed by random synthesis of a mixture of appropriate codon pairs and inserted into a bacterial plasmid in E. coli. Two of the isolated fragments were subjected to DNA sequence analysis and theoretically code for polypeptides containing up to 23% lysine, 12% tryptophan, 12% methionine, 6% isoleucine, and 6% threonine. These five amino acids make up 60% of the total amino acid content of the peptide, compared with 25% for the same amino acids in lactalbumin, a milk protein considered to be high in essential amino acids. These fragments, when fused to an active bacterial promoter, which directs the synthesis of chloramphenicol acetyl transferase (CAT), cause bacteria, harboring these modified genes, to take up more lysine as compared to control cells and produce commensurately larger CAT polypeptides. This method of gene synthesis may permit production of polypeptides with a specified amino acid composition to supplement specific diets low in the essential amino acids.     

Endocytosis and degradation of native, cathepsin D-degraded, and glutathione-inactivated aldolase by perfused rat liver

The uptake and degradation of 125I-labeled (a) native aldolase, (b) cathepsin D-inactivated aldolase, and (c) aldolase inactivated by oxidized glutathione were studied in perfused rat liver. All three forms of aldolase were removed from the perfusion medium and degraded by the liver, but the uptake of the glutathione-inactivated enzyme (half-life in perfusate = 10 min) was much faster than that of the native enzyme (half-life = 30 min) or the cathepsin-inactivated enzyme (half-life = 42 min). The degradation of the enzyme was almost totally inhibited by leupeptin, indicating that thiol proteinases in lysosomes play an important role in the digestion process. Degradation of native and cathepsin D-inactivated aldolase appeared to be slower than that of the glutathione-inactivated enzyme but studies in which liver was preloaded with aldolase by perfusion at 19 degrees C and then warming to 37 degrees C indicated that the rate of degradation of all three forms was similar. It is concluded that the liver is capable of distinguishing between the glutathione-altered aldolase and native or partially degraded aldolase with regard to endocytosis, but that all three forms are degraded at similar rates once within lysosomes.     

Microbiological Contamination of Hospital Air: I. Quantitative Studies

The levels of airborne contamination in various areas of two hospitals were determined during a 15-month sampling period, using Casella and Andersen volumetric samplers. Based on nearly 5,000 samples, the mean count per ft³ ranged from 4.5 in obstetric-gynecology delivery rooms to 72.4 in waste-handling areas. The mean count for the entire hospital environment was on the order of 20 contaminants per ft³; 48% were associated with particles >5 μ diam, 30% with particles between 2 and 6 μ diam, and 22% with particles <2 μ diam. The airborne contamination was influenced by traffic, activity, ventilation considerations, and gross surface contamination, but not markedly by seasonal changes. When suitable control measures were implemented, the level of contamination could be diminished and kept low.     

Microbiological Contamination of Hospital Air: II. Qualitative Studies

Over 10,000 airborne microorganisms, isolated from various areas of two hospitals, were characterized according to colonial and microscopic morphology and certain physiological reactions, including penicillin resistance and hemolysis. On the basis of all isolates examined during a 15-month period, 42.6% were gram-positive cocci, 19.2% were gram-positive rods, 14.0% were gram-negative rods, 17.1% were molds, 2.2% were actinomycetes, 1.2% were yeasts, and the remainder were assorted diphtheroids and coccobacillary types. The distribution of types varied according to hospital area, locations within a given area, and level of gross airborne contamination, but did not vary significantly with season of the year. There appeared to be some relationship between contaminant particle size and type of organism associated with the particle. Distribution of penicillin-resistant types differed markedly in different hospital areas, with proportions ranging from 21.4% in surgery areas to 4.3% in incinerator rooms. Of all gram-positive cocci isolated, 34.9% were hemolytic, and 16.4% were penicillin-resistant.     

3-Hydroxypropyl amide formation from 1-aminocyclopropane-1-carboxylic acid by a cell-free ethylene-forming system from olive leaves

The low ethylene yield in a cell-free ethylene-forming system from olive tree leaves ( Olea europaea L. cv. Picual) was investigated. During the incubation, 1-aminocyclopropane-1-carboxylic acid (ACC) was extensively transformed into 3-hydroxypropyl amide (HPA). Enzyme extract, Mn²⁺ and oxygen are responsible for this reaction. Horseradish peroxidase (EC 1.11.1.7) can substitute for the enzyme extract in this reaction. HPA formation could be one reason for the poor in vitro conversion efficiency of ACC to ethylene.     

Thyrotropin-releasing hormone stimulates intestinal transit in young rats

The effect of intracisternal injection of thyrotropin-releasing hormone (TRH) on small intestinal transit of a charcoal bolus was investigated in 14-, 21-, 28- and 35-day-old and adult rats. Intracisternal TRH (15 micrograms in 2 microliters) was administered, and transit (distance traveled by the charcoal) was measured 120 min later. In all age groups, intracisternal TRH increased charcoal transit significantly (P less than 0.05) as compared to saline-treated controls. This increase in transit was not mimicked by intravascular TRH, and it was blocked in all age groups by prior intraperitoneal injection of atropine (2 micrograms/g body weight). Vagotomy blocked TRH-induced increases in small intestine transit in rats of 28 days and older. Prior intraperitoneal injection of the antiserotonin compound, cyproheptadine (1 microgram/g body weight) reduced TRH-induced increases in small intestine transit in all age groups. These results demonstrate that centrally administered TRH stimulates small intestine transit through both cholinergic and serotonergic mechanisms in rats as early as 14 days of age.