The effect of copper on frog skin. The role of sulphydryl groups

1. 1. Cu²⁺ at a concentration of 10⁻⁴ M, when applied to the external side of the frog skin produces an increase in the short-circuit current (I_sc).

2. 2. This effect was studied in skins of Rana temporaria adapted to cold (5°C) and room temperature (20°C), skins of Rana pipiens adapted to cold, and the results compared with those obtained previously with Rana ribibunda.

3. 3. The observed effect is less dependent upon the adaptation to cold than upon the functional state of the skin: skins with low short circuit currents have a bigger response to Cu²⁺ than skins with high I_sc.

4. 4. A species difference cannot be ruled out since skins of Rana ribibunda exhibiting high I_sc give good responses to Cu²⁺.

5. 5. 5,5′-dithiobis(2-nitrobenzoic acid), a sulphydryl-oxidizing reagent, produces an effect similar to that of Cu²⁺, and dithiothreitol an SH-reducing agent, reverses the effect of this ion.

6. 6. Cu²⁺ also induces an increase in the unidirectional K⁺ fluxes and unmasks a net outward potassium flux.

7. 7. The outward K⁺ flux induced by Cu²⁺ is sensitive to ouabain.

8. 8. It is concluded that Cu²⁺ increases the permeability of the external barrier of the frog skin to Na⁺ and K⁺, probably by reacting with SH groups.

Isozymes of glutamate dehydrogenase from oat leaves: Properties and light effect on synthesis

Ammonium-dependent induction of a GDH isozyme in oat leaveswas proportional to light intensity and inhibited by DCMU. Thestimulation of GDH synthesis in response to ammonia was partiallyrepressed by benzimidazole. The inducible (no. 1) and noninducible(no. 2) GDH isozymes wereseparated and purified by approximately54 and 24 fold respectively. The two isozymes were highly specificfor NAD and the rate of NADH oxidation was 7 to 9 times higherthan NAD reduction. Both isozymes showed similar Km values forsubstrates of the reductive amination reaction and pH optimafor NADH oxidation. The pH optima for NAD reduction were 9 and8.2 for isozymes1 and 2 respectively. The two isozymes had asimilar molecular weight, 2.2–2.4 x 10⁵ but differed intheir isoelectric point and temperature sensitivity. Resultssuggest that the GDH isozymes in oat leaves are two differententities but might possess a similar metabolic function. (Received January 6, 1976; )     

Variable-volume continuous cultivation

Equations are developed which describe variable-volume cultivations, including fed-batch systems. An analogy is drawn between the quasi-steady state in variable-volume cultivation and a dynamic steady state in variable-flow, constant-volume chemostat bioreactors. Switching procedures are developed to give a steady-state transition from batch to fed-batch and to continuous operation. In this respect, considerations in the literature have been extended. Computer solutions of the governing differential equations verify the theory and provide insight into the behavior of variable-volume stirred tank reactors. Application of variable-volume cultivation as a tool in investigating growth rates at low substrate levels is suggested. Variable-volume bioreactor systems could be also to obtain controlled dynamic conditions for research or production purposes.     

Spontaneous endometrial tumours of 'Sabra' rats

In a 2 year study of 171 female (101 virgin; 70 multiparous) 'Sabra' rats, spontaneous endometrial tumours were found in 69% of 2 year old animals. Tumour development appeared to be age related, and only virgin females showed tumours before 18 months of age. Polyps were the most prevalent tumour type, followed by adenocarcinomas. The Sabra rat can be included among those rat strains having a high incidence of spontaneous endometrial neoplasia.     

Incorporating physiology into species distribution models moderates the projected impact of warming on selected Mediterranean marine species

Species distribution models (SDMs) correlate species occurrences with environmental predictors, and can be used to forecast distributions under future climates. SDMs have been criticized for not explicitly including the physiological processes underlying the species response to the environment. Recently, new methods have been suggested to combine SDMs with physiological estimates of performance (physiology-SDMs). In this study, we compare SDM and physiology-SDM predictions for select marine species in the Mediterranean Sea, a region subjected to exceptionally rapid climate change. We focused on six species and created physiology-SDMs that incorporate physiological thermal performance curves from experimental data with species occurrence records. We then contrasted projections of SDMs and physiology-SDMs under future climate (year 2100) for the entire Mediterranean Sea, and particularly the ‘warm’ trailing edge in the Levant region. Across the Mediterranean, we found cross-validation model performance to be similar for regular SDMs and physiology-SDMs. However, we also show that for around half the species the physiology-SDMs substantially outperform regular SDM in the warm Levant. Moreover, for all species the uncertainty associated with the coefficients estimated from the physiology-SDMs were much lower than in the regular SDMs. Under future climate, we find that both SDMs and physiology-SDMs showed similar patterns, with species predicted to shift their distribution north-west in accordance with warming sea temperatures. However, for the physiology-SDMs predicted distributional changes are more moderate than those predicted by regular SDMs. We conclude, that while physiology-SDM predictions generally agree with the regular SDMs, incorporation of the physiological data led to less extreme range shift forecasts. The results suggest that climate-induced range shifts may be less drastic than previously predicted, and thus most species are unlikely to completely disappear with warming climate. Taken together, the findings emphasize that physiological experimental data can provide valuable supplemental information to predict range shifts of marine species.     

Arginine substitution by alanine at the P1 position increases the selectivity of CmPI-II,a non-classical Kazal inhibitor

CmPI-II is a Kazal-type tight-binding inhibitor isolated from the Caribbean snail Cenchritis muricatus. This inhibitor has an unusual specificity in the Kazal family, as it can inhibit subtilisin A (SUBTA), elastases and trypsin. An alanine in CmPI-II P1 site could avoid trypsin inhibition while improving/maintaining SUBTA and elastases inhibition. Thus, an alanine mutant of this position (rCmPI-II R12A) was obtained by site-directed mutagenesis. The gene cmpiR12A was expressed in P. pastoris KM71H yeast. The recombinant protein (rCmPI-II R12A) was purified by the combination of two ionic exchange chromatography (1:cationic, 2 anionic) followed by and size exclusion chromatography. The N-terminal sequence obtained as well as the experimental molecular weight allowed verifying the identity of the recombinant protein, while the correct folding was confirmed by CD experiments. rCmPI-II R12A shows a slightly increase in potency against SUBTA and elastases. The alanine substitution at P1 site on CmPI-II abolishes the trypsin inhibition, confirming the relevance of an arginine residue at P1 site in CmPI-II for trypsin inhibition and leading to a molecule with more potentialities in biomedicine.