Biotoxic activity in the Mucorales

The toxigenicity of representatives of 15 species of Mucorales (Absidia glauca, Actinomucor elegans, Cunninghamella elegans, Helicostylum piriforme, Mortierella isabellina, Mortierella (Mucor) rammaniana, Mucor hiemalis, Mucor mucedo, Mucor spinosus, Phycomyces blakesleeanus, Rhizopus oligosporus, Rhizopus stolonifer, Syncephalastrum racemosum, Thamnidium elegans, Zygorhynchus moelleri) towards the larvae of brine shrimp (Artemia salina) and the growth of pea seedlings (Pisum sativum) and tobacco plants (Nicotiana tabacum) was evaluated. The fungi were cultivated on malt extract agar and aqueous solutions of the cultures were tested.Thamnidium elegans showed a marked toxic action towards brine shrimp (mortality: 74.1%) andPhycomyces, Actinomucor andSyncephalastrum were only weakly toxic. Length and weight of stems of pea seedlings were moderately reduced by extracts ofAbsidia, Cunninghamella, Zygorhynchus andThamnidium and to a lesser degree byMucor spinosus. Cunninghamella andMucor spinosus also inhibited the development of pea hypocotyls. The length of tobacco stems was reduced byMortierella ramanniana, Rhizopus stolonifer andCunninghamella elegans. Wilting or other toxic phenomena were never observed with both test plants. Considering the present results and data from literature it is suggested that species of Mucorales have only a weak toxigenicity.     

Pitfalls in the assay of carboxymethylcellulase activity

A purified endocellulase from Sclerotium rolfsii and a crude cellulase preparation from Trichoderma reesei are used to illustrate several pitfalls associated with the assay of carboxymethylcellulase activity and the subsequent attainment of linear enzyme dilution curves. It is shown that the nature of both the enzymes and the substrate make the assay unsuitable for use in the calculation of enzyme recovery and purity.     

Carbonic anhydrase activity in peripheral T-lymphocytes and appearance of the activity during their maturation in the thymus

Summary Carbonic anhydrase (CA) activity is demonstrated in lymphoid tissue for the first time using the histochemical (Hansson's) method. A CA-positive reaction was seen in lymphocytes present in T-lymphocyte areas in both the lymph node and the spleen. The most intense staining was seen in the small T-lymphocytes, whereas the mediumsized T-lymphocytes were less markedly stained. The cortical lymphocytes in the thymus were completely devoid of staining, but the small medullary T-lymphocytes stained intensely. The results suggest that peripheral and thymic medullary T-lymphocytes contain CA activity, which appears in these cells during their maturation in the thymus.     

Tryptophan 110, a residue involved in the toxic activity but not in the enzymatic activity of notexin

We prepared two derivatives of notexin, a phospholipase A2 from Notechis scutatus scutatus venom, by modifying the protein with 2-nitrophenylsulfenylchloride, a tryptophan-specific reagent. One derivative was modified at both tryptophans 20 and 110 whereas the other was modified at tryptophan 20. Evidence based on circular dichroic analysis and antigenicity towards a notexin-specific monoclonal antibody indicated that derivatization at both tryptophans did not affect the tertiary structure of notexin. Concomitant modification of tryptophans 20 and 110 induced a marked decrease in the capacity of notexin to kill mice and to block neuromuscular transmission in the chick biventer cervicis preparation, whereas selective modification at tryptophan 20 had no effect on the lethal properties of notexin. This implies that the decrease in the lethal properties of notexin after derivatization was due to modification at tryptophan 110. However, the diderivatized notexin retained full enzymatic activity, implying that neither tryptophan 20 and tryptophan 110 are involved in the catalytic function of the molecule. We conclude that notexin harbours two functional sites. One of them corresponds to the enzymatic site, whereas the other, which includes tryptophan 110, provides specific toxic characteristics to notexin. By reference to previous crystallographic studies, the relative spatial positions of elements involved in toxicity and the catalytic site, we propose a possible orientation of notexin with respect to its putative membrane-bound target.