Trichoderma reesei sequences that bind to the nuclear matrix enhance transformation frequency

Three DNA fragments, trs1, 2 and 3, were isolated from the Trichoderma reesei genome on the basis of their ability to promote autonomous replication of plasmids in Saccharomyces cerevisiae. Each trs element bound specifically to the isolated T. reesei nuclear matrix in vitro, and two of them bound in vivo, indicating that they are matrix attachment regions (MARs). A similar sequence previously isolated from Aspergillus nidulans (ans1) was also shown to bind specifically to the T. reesei nuclear matrix in vitro. The T. reesei MARs are AT-rich sequences containing 70%, 86% and 73% A+T over 2.9, 0.8 and 3.7 kb, respectively for trs1, 2 and 3. They exhibited no significant sequence homology, but were shown to contain a number of sequence motifs that occur frequently in many MARs identified in other eukaryotes. However, these motifs occurred as frequently in the trs elements as in randomly generated sequences with the same A+T content. trs1 and 3 were shown to be present as single copies in the T. reesei genome. The presence of the trs elements in transforming plasmids enhanced the frequency of integrative transformation of T. reesei up to five fold over plasmids without a trs. No evidence was obtained to suggest that the trs elements promoted efficient replication of plasmids in T. reseei. A mechanism for the enhancement of transformation frequency by the trs elements is proposed. Received: 1 March 1997 / Accepted: 13 May 1997     

High performance liquid chromatography,chemical laboratory practice: By Heinz Engelhardt. Springer-Verlag,New York/Berlin, 1979. 248 pp. $29.80

A gel filtration method has been developed for the complete removal of sodium dodecyl sulfate (SDS) from proteins and peptides. The protein or peptide (20 μg–10 mg) containing SDS (up to 30–60 mg) is dissolved in a mixture of propionic acid, formic acid, and water (2:1:2, $\text{v}\text{v}$). Under these conditions, protein-SDS (or peptide-SDS) complexes, as well as SDS micelles, are dissociated. Subsequently, protein and SDS can be separated on a small Sephadex G-25 superfine column. The recovery of protein is typically 90% or more.     

Improved assay of coenzyme F420 analogues from methanogenic bacteria

Summary An improved method for separating analogues of coenzyme F₄₂₀ by isocratic reversed-phase high performance liquid chromatography is described. The method offers improved resolution, shorter chromatography runs and requires less complex apparatus.     

Accelerated disulfide reduction with polyunsaturated fatty acids: a mechanism of ionic channel modulation?

Summary

Polyunsaturated fatty acids (PUFAs) have been shown to modulate the activity of ionic channels by an unknown mechanism. Some channels are activated (i.e. certain delayed-rectifier, potassium channels) and others are inhibited (i.e. certain calcium, sodium and other potassium channels). We have previously demonstrated that PUFAs can act as electron carriers. It is known that ionic channels can be redox modulated. The ability of fatty acids to serve as electron shuttling agents is proportional to their unsaturation. These PUFAs cause reduction of disulfides through a superoxide radical-independent mechanism, probably related to enhanced electron delocalization. The present study shows that there is a strong correlation between the ability of a PUFA to transfer an electron to a disulfide and its reported ability to modulate ionic channels. This suggests that electron transfer could be the mechanism of PUFAs action on particular ionic channels.     

Blastomycosis in wild wolves

Blastomycosis was fatal to a wild wolf in Minnesota, and serologic evidence of blastomycosis was found in a Wisconsin wolf. No unusual movements were detected in the Minnesota animal from October 1983 through October 1985. However, by early December 1985, this wolf was weak and debilitated, and it perished on 14 December after approaching a human residence.     

Delayed-onset synergism between leukotriene B4 and prostaglandin E2 in human skin

The time-course of cutaneous inflammatory responses to LTB4 and PGE2 both alone and in combination has been studied in 10 healthy volunteers. LTB4 induced a transient wheal and flare response in some subjects, maximal at 15 minutes and succeeded by an erythematous, indurated lesion at 2-4 hours. PGE2 elicited a wheal and erythema response which resolved within 1-2 hours. Combination of LTB4 and PGE2 produced acute wheal and erythema responses which did not differ significantly from the summation of responses to the individual constituents of the mixture or from responses to a two-fold increase in the concentration of either component. Wheal and erythema responses persisted, however, with significant potentiation of responses 4 hours after injection. As both leukotrienes and prostaglandins are generated in acute allergic reactions, the effects of these mediators in combination could contribute to persisting and late-onset responses to allergen, in both the skin and lung. In particular, sustained responses to the combination of LTB4 and PGE2 might be important in the pathogenesis of inflammatory skin diseases such as psoriasis.     

Hydroxylamine is a vasorelaxant and a possible intermediate in the oxidative conversion of L-arginine to nitric oxide

Our objective was to determine whether hydroxylamine is a possible intermediate in the oxidative conversion of L-arginine to nitric oxide. Vasorelaxation by hydroxylamine is known to be mediated by nitric oxide. The vasorelaxant properties of hydroxylamine were examined using rat aortic rings and an isolated rat lung perfusion model. Hydroxylamine and acetylcholine were equally effective in relaxing norepinephrine-contracted intact aortic rings, whereas only hydroxylamine relaxed aortic rings with endothelium removed. This endothelium-independent vasorelaxation by hydroxylamine indicated that the hydroxylamine-converting enzyme is not localized solely within endothelial cells. Catalase, an enzyme known to oxidize hydroxylamine to nitric oxide, was present in homogenates of intact and endothelium-denuded rings. Cyanamide, another catalase substrate and a known precursor of nitroxyl (HNO), was not a vasorelaxant of aortic rings or of isolated, hypoxia-constricted lungs. These results suggest that free nitroxyl is not an intermediate in the oxidation of hydroxylamine to nitric oxide. An overall pathway for the oxidative conversion of L-arginine through an hydroxylamine intermediate to nitric oxide is proposed.