Acta biochimica et biophysica; Academiae Scientiarum Hungaricae最新文献

Procine kidney aminoacylase (E.C. 3.5.1.14) is inhibited neither by phenylmethylsulfonylfluoride nor by alkylating agents (iodoacetate or iodoacetamide). Therefore reaction mechanisms including the formation of acylenzyme through seryl or cysteinyl side chains are ruled out. The enzyme is a metalloprotein that can be inactivated by ECTA and in which Co2+ is an equivalent substitute for the Zn2+ ion. The two SH groups/subunit of aminoacylase exhibit different reactivites to p-hydroxymercuribenzoate. Modification of the less reactive SH group reversibly inactivates the enzyme. We suggest that this cysteinyl side chain is situated in the active center or in its immediate vicinity. On the basis of our results we suppose a close similarity between aminoacylase and carboxypeptidase A with respect to their active center and catalytic mechanism.

The effect of polymixin, nigericin, gramicidin on the 42K-efflux of E. coli cells was studied. The 42K-efflux of the bacteria decreases in time according to an exponential function. The slopes of the linearized functions characterizing the efflux increase with increasing concentration of antibiotics. The frequency of events of the 42K-release as a parameter of antibiotics membrane interaction was determined on the basis of a theoretical model developed for the evaluation of our experimental data. In this way a quantitative comparison of the effectiveness of antibiotics was possible. We found, that the most effective antibiotic was polymixin, followed by nigericin and gramicidin.

The effect of immobilization for different periods (7, 14, 28 and 42 days) on the aerobic and anaerobic metabolism of rabbit muscles with different functions was studied by measuring the total activity of LDH and MDH as well as the distribution of their isoenzymes. The results showed that on plaster casting the characteristic aerobic metabolism of m. soleus declined with a concomitant increase in glycolytic processes. In m. gastrocnemius the anaerobic metabolism of the muscle was relatively decreased on immobilization, but glycolysis remained the prevailing energy yielding process. On immobilization the metabolism of m. soleus and that of m. gastrocnemius approached each other. This may indicate a certain dedifferentiation of the muscle tissue.

Gas chromatographic analysis of steroids using Sp-2100 stationary phase revealed that all urine samples from 34 healthy children (aged 4-14 years) and from 35 adults contained 5-pregnene-3 beta, 20 alpha-diol, which was also identified by GC-MS. The extent of pregnenediol excretion continuously increased from prepuberty to adult age. The observed increase in pregnenediol excretion during prepuberty points to an enhanced biosynthetic activity of the adrenal gland in the years preceding sexual maturation.

Some dynamic properties of protein- and antibody-treated cell membranes were investigated. The membrane viscosity of sheep erythrocytes increased upon binding various proteins as determined from the fluorescence emission anisotropy of 1,6-diphenyl 1,3,5-hexatriene dissolved in the membranes. However, specific, IgM type immunoglobulins produced against the erythrocytes decreased the intramembrane viscosity of cells. The applicability of the above dye to follow the binding of specific and non-specific proteins to erythrocytes is also discussed.

Triton X-100 and Lubrol PX at similar concentrations extracted more phospholipid than protein from rat liver and rabbit heart membranes, as compared with their original protein and phospholipid contents, respectively. All the types of membrane phospholipid were solubilized from rat liver membrane with the used non ionic detergents, but less sphingomyelin was extracted than phosphatidylcholine and phosphatidylethanolamine.

The K+ activated ATPase activity of myosin subfragment-1 was measured under different conditions and enzyme kinetic parameters were calculated. The logarithm of Km varies linearly with ionic strength down to very low KCl concentrations, the logarithm of k2 vs. ionic strength, on the other hand, considerably deviates from linearity at low concentrations of KCl. ADP is a competitive inhibitor, like myosin ATPase, and with practically the same inhibitor constant. The energetical parameters of the decomposition (to products and enzyme) of the S-1--ATP complex are partically the same as those for myosin, the parameters of its formation, however, differ from the corresponding values for myosin: delta SI is considerably, delta HI significantly higher in the case of myosin. This may be the result of some kind of interaction of the two heads of myosin.

1. The kinetic properties of pancreatic elastase (E.C. 3.4.21.11) were investigated with 33 tripeptidyl-p-nitroanilide substrates, and the Km, kcat and kcat/Km values were determined. 2. The individual contributions of the substrate side chains to the kinetic constants were evaluated by regression analysis. As a result of the additivity of the contributions, the kinetic parameters of any substrate constructed from the amino acid investigated can be predicted. 3. Suc-D-Phe-Pro-Ala-pNA was the best substrate, as far as the Km (Michaelis constant) is concerned. The compound was synthesized and assayed. An excellent correlation was observed between its calculated (14 microM) and experimentally determined (15 microM) Km values. The aldehyde derivatives of this substrate is a competitive inhibitor of elastase (Ki = 0.6 mM). 4. The contribution values of the best substrates permitted us to characterize the topography of subsites involved in the formation of the enzyme-substrate complex. This, in turn, led us to the conclusion that the S3-P3 interaction is relatively less important in the binding of good substrates.