N2O as a substrate and as a competitive inhibitor of nitrogenase

We have investigated the inhibitory effect of N2O on NH3 formation by purified component proteins from Klebsiella pneumoniae and have confirmed that the inhibition is competitive with respect to N2 and that N2O is reduced to N2, which in turn is further reduced to NH3. In addition, we have shown that N2O is unable to support HD formation from D2 and H2O. N2-supported HD formation from D2 and H2O was found to be inhibited by N2O. In contrast to N2, N2O was found to suppress nitrogenase-mediated H2 evolution completely at infinitely high pN2O. H2 was found to inhibit N2O-supported NH3 production but not N2O-supported N2 production. The steady-state kinetics of N2O reduction showed a good fit to Michaelis-Menten kinetics with a Km for N2O of 5 mM at 30 degrees C, corresponding to 24 kPa of N2O. A model is proposed that fits the observed results.     

Assessment of adult skeletons to detect prenatal exposure to trypan blue in mice

A Skeletal Variant Assay System (SVAS) consisting of a group of 88 spontaneously occurring qualitative variations of the adult mouse skeleton was studied in CD-1 mice which had been exposed in utero by way of three daily ip injections of their dams on days 7-9 of gestation with trypan blue. Treatment groups received daily doses of 0.25 cc of 0, .037, .075, .15, or .30% trypan blue dissolved in 0.9% NaCl. Two separate series of experiments were performed, and skeletons were examined at 62 +/- 2 days postnatal. Sixty-six and 58 of the variants occurred in the two series, respectively. Frequencies of occurrence of substantial numbers of variants differed from Untreated (UNTD) and Vehicle-Treated (VEH) values in a dose-related manner for both series. At the high dose 18 and 22 variants occurred with significantly different (P less than .01) frequencies from UNTD in the two series. Contrasting high-dose animals with vehicle controls revealed significant differences in 24 and 17 variants. There were 13 and 14 variants in the two series, respectively, which differed from both UNTD and VEH. If one considers differences at P less than .01 in one comparison and P less than .05 in the other, then 22 and 18 variants qualify as being significantly different from both controls in the two series. Agreement between the two series was excellent regarding which variants were affected. Several differed significantly from both UNTD and VEH in both series of experiments. Among these were a number which appeared more or less specific to trypan blue exposure. They include Dyssymphysis of the Atlas, Carpal Fusions, and Tarsal Fusions. Although increased frequency of an Interfrontal bone is seen with several treatments, the magnitude of the response and the low doses at which it is elicited are unique to trypan blue exposure. Numerous low-dose effects are striking in this set of experiments, making the SVAS a very sensitive indicator of trypan blue exposure. In addition to the variants mentioned, a large cluster of cervical (C) vertebrae variants, including dyssymphyses, fusions, imperfect transverse foramina of C1 and C2, and accessory transverse foramina of C3-C6, as well as vertebral fusions at various levels (especially cervical, sacral, and caudal), appear to be the principal effects of exposure to this compound. In addition, rib malformations at the high dose level, and increased frequency of occurrence of 27-presacral vertebrae at all dose levels, were important effects.(ABSTRACT TRUNCATED AT 400 WORDS)     

Apparent in vivo retroconversion of dietary arachidonic to linoleic acid in essential fatty acid-deficient rats

Essential fatty acid-deficient rats were fed ethyl [U-14C]arachidonate (308 dpm/nmol) and when a decrease in the transepidermal water loss was seen, the epidermal sphingolipids, acylglucosylceramide and acylceramide were isolated. [14C]Linoleic acid (approx. 130 dpm/nmol) was present in both lipid classes, while the substrate was only detected in the former. These results intimate that in vivo retroconversion of arachidonic to linoleic acid can be induced in the rat.     

The relative abundance and seawater requirements of gram-positive bacteria in near-shore tropical marine samples

The relative abundance of gram-positive bacteria in a variety of near-shore marine samples was determined using the KOH method. Gram-positive bacteria accounted for 14%, 25%, 31 %, and 12%, respectively, of the colony-forming bacteria obtained from seawater, sediments, and the surfaces of algae and invertebrates. A total of 481 gram-positive strains were isolated representing a wide range of morphological groups including regular and irregular rods, cocci, and actinomycetes. Seventy-seven percent of the strains characterized did not form spores and were aerobic, catalase-positive rods with regular to irregular cell morphologies. Eighty-two percent of the strains tested showed an obligate requirement of seawater for growth. None of the cocci tested required seawater or sodium for growth. This is the first report documenting that gram-positive bacteria can compose a large percentage of the culturable, heterotrophic bacteria associated with the surfaces of tropical marine algae. Correspondence to: P.R. Jensen     

Loss of allosteric control but retention of the bifunctional catalytic competence of a fusion protein formed by excision of 260 base pairs from the 3' terminus of pheA from Erwinia herbicola.

A bifunctional protein denoted as the P protein and encoded by pheA is widely present in purple gram-negative bacteria. This P protein carries catalytic domains that specify chorismate mutase (CM-P) and prephenate dehydratase. The instability of a recombinant plasmid carrying a pheA insert cloned from Erwinia herbicola resulted in a loss of 260 bp plus the TAA stop codon from the 3' terminus of pheA. The plasmid carrying the truncated pheA gene (denoted pheA*) was able to complement an Escherichia coli pheA auxotroph. pheA* was shown to be a chimera composed of the residual 5' part of pheA (901 bp) and a 5-bp fragment from the pUC18 vector. The new fusion protein (PheA*) retained both chorismate mutase and prephenate dehydratase activities. PheA* had a calculated subunit molecular weight of 33,574, in comparison to the 43,182-molecular-weight subunit size of PheA. The deletion did not affect the ability of PheA* to assume the native dimeric configuration of PheA. Both the CM-P and prephenate dehydratase components of PheA* were insensitive to L-phenylalanine inhibition, in contrast to the corresponding components of PheA. L-Phenylalanine protected both catalytic activities of PheA from thermal inactivation, and this protective effect of L-phenylalanine upon the PheA* activities was lost. PheA* was more stable than PheA to thermal inactivation; this was more pronounced for prephenate dehydratase than for CM-P. In the presence of dithiothreitol, the differential resistance of PheA* prephenate dehydratase to thermal inactivation was particularly striking.(ABSTRACT TRUNCATED AT 250 WORDS)     

Variance of ventilation during exercise.

Expired gas concentrations were measured during a multibreath washin of He in one female and seven male subjects at rest (seated) and during cycle exercise at work rates of 70-210 W. In a computational model, the ventilation distribution was represented as a log-normal distribution with standard deviation (sigmaV); values of sigmaV were obtained by fitting the output of the model to the data. At rest, sigmaV was 0.89 +/- 0.18; during exercise, sigmaV was 0.60 +/- 0.13, independent of the level of exercise. These values for the width of the functional ventilation distribution at the scale of the acinus are approximately two times larger than those obtained from anatomic measurements in animals at a scale of 1 cm3. The values for sigmaV, together with data from the literature on the width of the functional ventilation-perfusion distribution, show that ventilation and perfusion are highly correlated at rest, in agreement with anatomic data. The structural sources of nonuniform ventilation and perfusion and of the correlation between them are unknown.