Acta biochimica et biophysica; Academiae Scientiarum Hungaricae最新文献

Cold produced increase in latency of the dorsal root evoked monosynaptic ventral root reflex was investigated in frog lumbar spinal cord. Attempts were made to separate changes in the synaptic transmission time from changes in the conduction velocity of axons involved in the reflex pathway. Reduction of the temperature by 10 degrees C from the initial values of about 17-19 degrees C produced a 50-60 per cent increase in reflex latency, similar to the degree of slowing down of the axonal conduction velocity. The finding fails to conclusively support the chemical synaptic transmission mechanism of the ventral root discharges studied and calls for more detailed investigation of the phenomenon.

Hypotonic lysis was applied for the preparation of lymphocyte plasma membranes. By this method it was possible to separate membrane fractions with a high sialic acid to protein and cholesterol to phospholipid ratio. The plasma membrane fractions had a lower content of endoplasmatic reticulum than the preparations described earlier but were slightly contaminated with DNA. The isolation method proved to useful for the preparation of surface membranes from human lymphocytes

The authors determined the surface absorbed dose (Ds = 67-68 mGy) and the average absorbed dose (Dav = 13-15 mGy) by means of non-screen film radiography of the extremities. The authors recommend the use of xeroradiography (with a tungsten anode tube) or, first of all, a rare-earth screen - film combination (e.g. Kodak min-R + Nuklearmedizin NMB film). The two methods permit the dose values to be decreased by a factor of about 4 and 15, respectively. The dose values are smaller, at least for the 8 cm thick tissue equivalent phantom, when a tungsten anode is used instead of a Mo anode.

The superprecipitation of synthetic actomyosin, formed from intact myosin and actin extracted from rabbit white skeletal muscles 14 and 75 days after denervation has been compared with that of intact synthetic actomyosin. Superprecipitation has been characterized by two parameters. 1. The value of superprecipitation delta E determining the increase in the absorbance from minimum to maximum values; 2. the time required for the half-maximum increase (t 1/2) which is the inverse of the velocity constant. It has been shown that both the delta E and the velocity constant decrease: the delta E amounts to 75.4 +/- 7.3 per cent (p = 0.95) and 87.9 +/- 11.2 per cent (p = 0.95) after 14 and 75 days, respectively, when compared with normal; the t 1/2 increases to 41.5 +/- 13.6 per cent (p = 0.95) after 14 days and 17.5 +/- 11.9 per cent (p = 0.95) after 75 days. It is assumed that this effect is related to changes in the structure of actin.

Phosphorylase kinase from rabbit skeletal muscle inhibited the dephosphorylation of phosphorylase a by phosphoprotein phosphatase. Phosphorylation (activation) of phosphorylase kinase by cyclic AMP-dependent protein kinase greatly increased this inhibitory effect. Thus, phosphoprotein phosphatase is inhibited by phosphorylase kinase in a reversible manner (Gergely et al. (1976) Biochim. Biophys. Acta 429 809-816). In this paper the regulation by phosphorylase kinase at phosphoprotein phosphatase activity in different fractions of muscle extract and in the presence of various ligands has been investigated. The presence of phosphorylase kinase also affected the ligand control of phosphatase activity. Phosphorylase kinase almost cancelled the inhibitory effect of AMP but hardly influenced the activating effect of glucose, glucose 6-phosphate and caffeine. Calmodulin, glycogen and phosphorylase b (effectors of phosphorylase kinase) did not influence the inhibitory effect of phosphorylase kinase. Fractions of muscle extract also demonstrated the regulatory role of phosphorylase kinase. These fractions contained considerable amounts of phosphorylase kinase and phosphatase. Phosphatase activity was inhibited by phosphorylation reactions triggered by Mg++ and ATP. Heat-stable inhibitors were absent from these fractions, therefore the transient inhibition of phosphatase could be attributed to the phosphorylation of endogenous phosphorylase kinase. The introduction between phosphorylase kinase and phosphatase resulted in a loss of AMP sensitivity, i.e. AMP did not inhibit the activity of phosphatase in those fractions. Our results imply that the phosphorylation of phosphorylase kinase is equally important both in the formation of enzymatically active phosphorylase a and in the inhibition of dephosphorylation of phosphorylase a. The consequence of these two effects is the elevated level of phosphorylase a.

The interaction of rose bengal (RB) with rabbit skeletal muscle phosphorylase b (1,4-alpha-D glucan: orthophosphate alpha-glucosyl-transferase, E.C. 2.4.1.1.) was studied by kinetic and absorption photometric methods. RB inhibited the phosphorylase b activity. Inhibition was strictly competitive with respect to substrate G-1-P and activator AMP with inhibition constants 2 x 10(-6) M and 2.2. x 10(-7) M, respectively. The association of the dye with the enzyme elicited a red shift in the spectrum of RB indicating an apolar binding site. According to difference absorption measurements, the enzyme binds two dye molecules per dimer in the presence and absence of both G-1-P and AMP. Binding constants determined from photometric titrations are consistent with those obtained from kinetic measurements. The present findings allow to carry out detailed kinetic investigations on the activator AMP and substrate G-1-P binding of phosphorylase b.

A component was observed in the steroid spectrum of the urine of salt-loosing children with C/21-hydroxylase deficiency, which was eluted from Sp 2100 stationary phase before pregnanetriol but, unlike pregnanetriol, exhibited heat and acid stability. This component was isolated by paper chromatography and identified as 3 alpha, 20 alpha-dihydroxy-5 beta-pregnan-11-one by GC-MS and further gas chromatographic analysis. The amount of the steroid was minimal in the urine of infants, while in children submitted to substitution corticoid therapy it showed an increasing tendency, especially during puberty. The maximal value of excretion, in one case, amounted to 17% relative to total steroids. In puberty a significant excretion of 11-keto-pregnanediol indicates that under the given conditions the 11 beta-hydroxylation of steroid intermediates in the adrenals may be considerable not only at the level of 11-hydroxy-progesterone but also at that of progesterone.

Messenger ribonucleoprotein (mRNP) was released from 0.5 M KCl washed rat liver polyribosomes after mild pancreatic ribonuclease (EC 3.1.27.5) and EDTA treatment and separated by sucrose gradient centrifugation from ribosomal subunits. The method yielded partially fragmented mRNP, which, however, was free from ribosomal contaminants. In CsCl gradient the mRNP banded at 1.46 g/cm3, indicating a protein content of about 65%. Treatment of mRNP with 0.25 M or 0.5 M KCl resulted in loss of the proteins. Urea/sodium dodecyl sulfate polyacrylamide gel electrophoresis of mRNA bound proteins showed that the most prominent polypeptides found in the mRNP fractions exhibited molecular weights of 29 000 (P29), 31 000 (P31), 38 000 (P38), 44 000 (P44), 50 000 (P50), 54 000 (P54), 63 000 (P63) 76 000 (P76) and 105 000 (P105). Three polypeptides, P38, P44 and P63 were most sensitive to high salt treatment.

To elucidate some ambiguous details in the tryptic fragmentation scheme of HMM as given by Bálint et al. (J. Biol. Chem. 250 (1975) 6168; Arch. Biochem. Biophys. 190 (1978)793), the peptide fragments were isolated by a milligram scale preparative gel electrophoresis procedure. The dansyl-peptide map of the 20 kDal tryptic fragment obtained from tryptic heavy meromyosin (HMM) and that of a similar fragment from papainic subfragment-1 (S-1) were found to be nearly identical. This finding gives unequivocal proof of the location of the 17 kDal peptide stretch lost during digestion in the form of small peptides, at the C terminal part of the heavy chain backbone of HMM. The N terminals of the 150, 74, and 25 kDal fragments of the heavy chain isolated from HMM digested by trypsin under widely differing conditions were shown to be acetylated. The N terminal amino group of the other peptide fragments of HMM remains the same under widely differing conditions of digestion. We conclude that all the fragments are well defined polypeptides and digestion progresses by splitting from the C terminals formed by the primary splits.