Adenylate energy charge in streambed sediments

SUMMARY.

• 1 The adenylate energy charge (EC_A) of microbial communities from streambed sediments was measured during three different seasons, under experimental manipulation and in culture.
• 2 The EC_A values of sediments (x±S.E.) in the autumn, winter and spring were low and constant; 0.22±0.03 (n=12), 0.32±0.04 (n= 12) and 0.28±0.03 (n=6) respectively.
• 3 A 5 h exposure of sediments to an algal lysate at 3.0–4.0°C and a 48 h exposure of sediments to tryptone-yeast extract at 8.0–18.0°C failed to increase EC_A even though respiration increased 3.7-fold during the latter exposure.
• 4 The cellular EC_A of a bacterial monoculture, sampled in log phase, was 0.90±0.10 (n=3), but cxtracclluhir AMP depressed the total culture EC_A to 0.21±0.01 (n=4).
• 5 Attempts to isolate extracellular AMP from the interstitial waters of sediments were unsuccessful.
• 6 The data suggest that under natural conditions EC_A is of limited use as a monitor of subtle changes in the physiological state of microbial communities in streambed sediments.

     

A highly conserved nuclear gene for low-level phylogenetics: elongation factor-1 alpha recovers morphology-based tree for heliothine moths

Molecular systematists need increased access to nuclear genes. Highly conserved, low copy number protein-encoding nuclear genes have attractive features for phylogenetic inference but have heretofore been applied mostly to very ancient divergences. By virtue of their synonymous substitutions, such genes should contain a wealth of information about lower-level taxonomic relationships as well, with the advantage that amino acid conservatism makes both alignment and primer definition straightforward. We tested this postulate for the elongation factor-1 alpha (EF-1 alpha) gene in the noctuid moth subfamily Heliothinae, which has probably diversified since the middle Tertiary. We sequenced 1,240 bp in 18 taxa representing heliothine groupings strongly supported by previous morphological and allozyme studies. The single most parsimonious gene tree and the neighbor-joining tree for all nucleotides show almost complete concordance with the morphological tree. Homoplasy and pairwise divergence levels are low, transition/transversion ratios are high, and phylogenetic information is spread evenly across gene regions. The EF-1 alpha gene and presumably other highly conserved genes hold much promise for phylogenetics of Tertiary age eukaryote groups.      

Chick embryo fibroblasts produce two forms of hyaluronidase

Cultured chick embryo fibroblasts derived from skin and skeletal muscle exhibit hyaluronidase activity both associated with the cell layer and secreted into the medium. Although both forms of the enzyme have a number of similar characteristics (R.W. Orkin and B.P. Toole, 1980, J. Biol. CHem. 255), they differ in thermal stability at neutral pH and in behavior on ion-exchange chromatography. Both forms of the enzyme are equally stable at acidic pH for long intervals, but the cell-associated hyaluronidase is significantly less stable than the secreted froms at neutral pH and at temperatures more than or equal to 30 degrees C. Neither the presence of proteases nor inhibitors of hyaluronidase appear to be involved in the cell-asspcoated enzyme. Chromatography of the two forms of hyaluronidase on carboxymethyl cellulose reveals that most (60-90 percent) of the secreted form of the enzyme elutes at a lower ionic strength than the cell- associated enzyme. Treatment of the secreted form of hyaluronidase with neuraminidase shifts its elution profile on carboxymethyl cellulose toward that of the cell-associated form, and also decreases its thermal stability at neutral pH. In contrast, treatment of the secreted form of hyaluronidase with alkaline phosphatase has no detectable effect. These data suggest that the secreted hyaluronidase differs from the cellular form in possessing additional sialic acid residues which endow the former with increased stability in the extracellular milieu.     

The action of ultraviolet radiation on yeast catalase

The effect of prolonged UV irradiation (mostly 2537 A) on the catalase activity of an aqueous yeast suspension was divisible into 4 periods. First, the period during which the cells lost their ability to form colonies, but during which no change in catalase activity was noted. Second, the period during which a considerable rise in catalase activity (Euler effect) occurred. The Euler effect was accompanied by enzyme alteration as shown by the simultaneous decrease in the activation energy of the enzyme-substrate system. However, during the initial phase of this period, as the catalase activity of the suspension began to increase, the activation energy rose to a transient level higher even than that characterizing the unaltered enzyme. Heat accelerated the rate of alteration when applied either during or after the irradiation; the activation energy for the over-all alteration reaction was 24 kcal., a value close to that recorded previously for alteration induced by chemical agents. Nevertheless, the rate-limiting step appeared to be different in the two cases. A model of these events was presented in which the primary photochemical action was on the site at which catalase is located within the cell. Third, a rather long period during which irradiation led to no diminution in the catalase activity of the maximally active suspension. This protection effect was duplicated in intro by a model crystalline catalase-KNA system, or by adding either ribonuclease digestion products of RNA or adenine to a catalase solution prior to irradiation. Evidence was adduced that the protection effect was not a simple screening, but involved some sort of interaction between the enzyme and the nitrogenous components of RNA, an interaction which must likewise occur within the cell. Alteration induced by CHCl₃ did not eliminate the protection effect, but that by butanol did. The onset of photoinactivation was due to modification of protein structure, not of RNA. Fourth, the period of photoinactivation of the intracellular enzyme, which was quite similar to that of the crystalline enzyme in vitro.