On the mechanism of enzyme action. LIII. The amphoteric properties of trypsin

Several methods were employed for the preparation of salt-free trypsin samples which were used to determine the electrometric titration curves of the enzyme. These curves point to a maximum acid-binding capacity below pH 2. Stoichiometric analysis indicates the presence of 6 carboxyl groups per 10,000 g. of proteins, 1 imidazole group, and 13 hydroxyl-binding groups. Calcium has a specific effect on the titration curves by increasing the acidity of the carboxyl groups in the pH range 3.5 – 5. This effect is not shown by potassium, sodium, or even the bivalent magnesium ion. It is attributed to the formation of a specific complex between the enzyme and the calcium ions, involving the carboxyl groups of the protein. The equally specific protective effect of calcium on the self-digestion of trypsin can therefore be explained by assuming the formation of a complex which stabilizes the enzyme.     

Chronic exposure of diesel exhaust particles induces alveolar enlargement in mice

Background

Diesel exhaust particles (DEPs) are deposited into the respiratory tract and are thought to be a risk factor for the development of diseases of the respiratory system. In healthy individuals, the timing and mechanisms of respiratory tract injuries caused by chronic exposure to air pollution remain to be clarified.

Methods

We evaluated the effects of chronic exposure to DEP at doses below those found in a typical bus corridor in Sao Paulo (150 μg/m³). Male BALB/c mice were divided into mice receiving a nasal instillation: saline (saline; n = 30) and 30 μg/10 μL of DEP (DEP; n = 30). Nasal instillations were performed five days a week, over a period of 90 days. Bronchoalveolar lavage (BAL) was performed, and the concentrations of interleukin (IL)-4, IL-10, IL-13 and interferon-gamma (INF-γ) were determined by ELISA-immunoassay. Assessment of respiratory mechanics was performed. The gene expression of Muc5ac in lung was evaluated by RT-PCR. The presence of IL-13, MAC2+ macrophages, CD3+, CD4+, CD8+ T cells and CD20+ B cells in tissues was analysed by immunohistochemistry. Bronchial thickness and the collagen/elastic fibers density were evaluated by morphometry. We measured the mean linear intercept (Lm), a measure of alveolar distension, and the mean airspace diameter (D0) and statistical distribution (D2).

Results

DEP decreased IFN-γ levels in BAL (p = 0.03), but did not significantly alter IL-4, IL-10 and IL-13 levels. MAC2+ macrophage, CD4+ T cell and CD20+ B cell numbers were not altered; however, numbers of CD3+ T cells (p ≤ 0.001) and CD8+ T cells (p ≤ 0.001) increased in the parenchyma. Although IL-13 (p = 0.008) expression decreased in the bronchiolar epithelium, Muc5ac gene expression was not altered in the lung of DEP-exposed animals. Although respiratory mechanics, elastic and collagen density were not modified, the mean linear intercept (Lm) was increased in the DEP-exposed animals (p ≤ 0.001), and the index D2 was statistically different (p = 0.038) from the control animals.

Conclusion

Our data suggest that nasal instillation of low doses of DEP over a period of 90 days results in alveolar enlargement in the pulmonary parenchyma of healthy mice.

Electronic supplementary material

The online version of this article (doi:10.1186/s12931-015-0172-z) contains supplementary material, which is available to authorized users.     

Biosynthesis of chlorophyll P-680 of photosystem II in greening leaves of plants adapted to heat shock

In etiolated pea and maize leaves illuminated after incubation at 38 degreesC, a new dark reaction was shown manifested in the bathochromic shift of spectral bands and accompanied by esterification of the product of protochlorophyllide photochemical reduction--Chld 684/676: Chld 684/676 --> Chl 688/680. After completion of the reaction a rapid (20-30 sec) quenching of the fluorescence of the reaction product (Chl 688/680) was observed. The reaction Chld 684/676 --> Chl 688/680 is inhibited under anaerobic conditions and in the presence of cyanide; the reaction accompanied by Chl 688/680 fluorescence quenching is not observed in pea mutants with impaired function of photosystem II reaction centers. The spectral properties of the formed Chl form with the absorption maximum at 680 nm, fluorescence quenching, and simultaneous synthesis of pheophytin suggest that the reaction is connected with the chlorophyll of photosystem II reaction center--P-680.