Cloning, isolation, and characterization of mammalian legumain, an asparaginyl endopeptidase.

Legumain is a cysteine endopeptidase that shows strict specificity for hydrolysis of asparaginyl bonds. The enzyme belongs to peptidase family C13, and is thus unrelated to the better known cysteine peptidases of the papain family, C1 (Rawlings, N. D., and Barrett, A. J. (1994) Methods Enzymol. 244, 461-486). To date, legumain has been described only from plants and a blood fluke, Schistosoma mansoni. We now show that legumain is present in mammals. We have cloned and sequenced human legumain and part of pig legumain. We have also purified legumain to homogeneity (2200-fold, 8% yield) from pig kidney. The mammalian sequences are clearly homologous with legumains from non-mammalian species. Pig legumain is a glycoprotein of about 34 kDa, decreasing to 31 kDa on deglycosylation. It is an asparaginyl endopeptidase, hydrolyzing Z-Ala-Ala-Asn-7-(4-methyl)coumarylamide and benzoyl-Asn-p-nitroanilide. Maximal activity is seen at pH 5.8 under normal assay conditions, and the enzyme is irreversibly denatured at pH 7 and above. Mammalian legumain is a cysteine endopeptidase, inhibited by iodoacetamide and maleimides, but unaffected by compound E64 (trans-epoxysuccinyl-L-leucylamido-(4-guanidino)butane). It is inhibited by ovocystatin (cystatin from chicken egg white) and human cystatin C with Ki values < 5 nM. We discuss the significance of the discovery of a cysteine endopeptidase of a new family and distinctive specificity in man and other mammals.

Measurement, analysis, and modelling of the caloric response. 2. Evaluation of mental alerting tasks for measurement of caloric-induced nystagmus.

Management of the patient's level of arousal is one of the most important variables in obtaining consistent and strong caloric responses. The patient may suppress the caloric response and/or exacerbate beat-to-beat variability if some type of mental alerting (MA) task is not used to focus the patient's attention from his or her dizziness. An experiment was undertaken to evaluate simple MA tasks in terms of associated caloric response strength and variability. Warm caloric responses were measured while each of 10 normal subjects performed eight different MA tasks. The mental exercises included two math tasks, two quizzing tasks, two hand-motor tasks, and two alphabet tasks. One of the tasks in each complementary pair required the subject to interact with the examiner throughout the caloric response. Minimal or no interaction was required for the companion task. The relative ordering among the eight MA tasks was compared in terms of total sum of ranks, summed across 15 performance measures taken from caloric response indices. The highest-ranked altering task was an exercise requiring subjects to name or list local cities, states in the U.S.A., colors, etc. The lowest-ranked tasks were backward counting exercises and reflexive quizzing, which are the traditional tasks used in the clinic.

Monocular and binocular visual evoked potentials before and after cataract surgery.

Visual evoked cortical potentials (VEPs) were recorded from an individual with a mature cataract in one eye. Stimulation was both monocular and binocular and VEPs were obtained from three occipital scalp sites. Comparisons of recordings taken prior to cataract removal and after removal showed a dramatic increase in amplitude of potentials derived through stimulation of the affected eye. Slight differences in hemispheric amplitude prior to surgery suggest a greater degree of opacity in one portion of the affected lens.

The effect of shearing pregnant ewes on lamb birth weights.

Shearing housed pregnant ewes at about the 11th week of pregnancy resulted in increased ewe liveweight gain (P greater than 0-01) and heavier twin litter weights (P greater than 0-01). The lambs from the shorn ewes also grew faster when put out to pasture. Unshorn pregnant ewes had a faster and more variable respiration rate, probably due to their being too hot in the prevailing conditions. The effect of this apparent heat stress on fetal nutrition is discussed.