Fluctuations and mechanical strength of α-helices of polyglycine and poly(L-alanine)

By using the static correlations of fluctuations in the dihedral angles of the α-helices of polyglycine and poly(L -alanine) calculated previously, geometrical fluctuations of a section (consisting of up to 18 peptide units) of the α-helices of infinite length are calculated. These fluctuations are found to differ in some respects (e.g., the dependence of amplitudes on the length of section) from those of a circular rod made of homogeneous continuous material. However, the moduli of the mechanical strengths (tensile Young's modulus, bending Young's modulus, and the shear modulus) of a circular rod are calculated, whose geometrical fluctuations are approximately equal to the fluctuations of a section consisting of 18 peptide units. They are of the order of 10¹¹ dyn/cm². The tensile rigidity, flexural rigidity, and torsional rigidity are calculated to be 1.20 × 10^?3 dyn, 2.46 × 10^?19 dyn·cm² and 1.79 × 10^?19 dyn·cm² for polyglycine, and 1.96 × 10^?3 dyn, 4.05 × 10^?19 dyn·cm² and 3.28 × 10^?19 dyn·cm² for poly(L -alanine), respectively.     

Polymerization of amino acid derivatives by polymer catalysts. V. Polymerization of DL-β-phenylalanine N-carboxyanhydride by several poly(N-alkylamino acid) diethylamides

The polymerization of DL -β-phenylalanine N-carboxyanhydride (NCA) initiated by poly(N-benzylglycine)diethylamide (DEA) and poly(N-methyl-DL -alanine)DEA has been investigated. As previously reported, polysarcosine DEA, poly-N-ethylglycine DEA, and poly-N-n-propylglycine DEA showed marked accelerations in the polymerization of DL -β-phenylalanine NCA as compared with the polymerization initiated by low molecular weight, amines having similar base strength. However, this phenomenon (the chain effect) was not observed with the two polymer catalysts studied in the present investigation With poly-N-methyl-DL -alanine DEA, adsorption of DL -β-phenylalanine NCA onto the polymer chain takes place, though not so effectively as with other polypeptides, so the absence of chain effect was ascribed to a reduced flexibility of the polymer chain. With poly(N-benzylglycine)DEA, the reactivity of terminal base group was found to be much lower than that of other polymer catalysts. However, the absence of the chain effect would be attributed to the rigidity of polymer chain of poly-N-benzylglycine DEA due to the bulkiness of the N-benzyl group.     

Bacillus stearothermophilus KP 1236 neutral protease with strong thermostability comparable to thermolysin

Summary An extracellular neutral protease, of Bacillus stearothermophilus KP 1236 (a soil isolate) able to grow at 39°–71°C was purified to homogeneity. The molecular weight, sedimentation coefficient in water at 20°C, and isoelectric point were estimated as 33,000, 3.46 S and 7.5, respectively. The enzyme was most active at 80°C and pH 7.0. The activity was stable for 10 min up to 80°C at pH 7.5 and for 18 h at 60°C over pH 6.0–8.8. The enzyme and thermolysin (microbial metalloproteinase, EC 3.4.24.4) shared their antigenic determinants in part.Presented at the Annual Meeting of the Agricultural Chemical Society of Japan on Sapporo, July 30, 1985 (Abstracts, p. 333)     

Anti-tumour effect of humoral and cellular immunities mediated by a bacterial immunopotentiator,Lactobacillus casei,in mice

Summary Administration of a mixture containing Lactobacillus casei YIT 9018 (LC9018) and methylcholanthrene-induced fibrosarcoma (Meth A) cells into the peritoneum of syngeneic BALB/c mice suppressed the tumour growth and protected the mice from tumour death. With the appearance of the anti-tumour activity, serum complement-dependent tumour cytotoxic (CDC) antibody was induced on the 5th day after the administration as a result of the adjuvant effect. The cytotoxic antibody was not found in serum on the 5th day after inoculation of Meth A cells alone, but it was induced before the mice died of the tumours. Adjuvant induction of the cytotoxic serum antibody at an early time was also observed using Kirsten murine sarcoma virus-transformed tumour (K234) cells. Both of these cytotoxic antibodies in sera from Meth A-suppressed and the tumour-bearing mice were specific for the tumour cells and were IgM class, since they were absorbed with rabbit anti-mouse IgM antibody. However, the cytotoxic antibody was not found in the peritoneal cavity which was the tumour inoculation site, but binding antibody against the tumour cells was faintly detected in the region using an enzyme-linked immunoabsorbent assay (ELISA). In neutralization tests, the cytotoxic antibody did not exert anti-tumour activity in recipient mice when it was administered to the mice along with the tumour cells or when it was administered i. v. at the time of tumour inoculation. Moreover, the cytotoxic antibody was not available for the antibody-dependent cell-mediated cytotoxicity (ADCC). These results suggest that the cytotoxic antibody did not exert anti-tumour activity in the tumour-suppressed mice. In contrast, peritoneal exudate cells (PEC) on the 5th day, and PEC and spleen cells on the 15th day after i. p. administration of the mixture exerted strong anti-tumour activity as measured by the Winn test.In conclusion, the adjuvant effect of LC9018 induced tumour-specific humoral and cellular immunities but the anti-tumour activity was dependent only on the cellular effectors of the host. The possible use of LC9018 in tumour immunotherapy is discussed.     

Solvent effect in the stereoselective polymerization of α-amino acid N-carboxyanhydride and some considerations on the mechanism of stereoregulation

In the Polymerization of phenylalanine N-carboxyanhydride (NCA) in No₂Oh initiated by MeNHBzl, L -,D -, and DL -NCA As were polymerized at the same rate, and no stereoselectivity was observed. When the same experiment was carried out in HCONEt₂, however, L - and D -NCA were both polymerized at a rate which was about twice as large as that of DL -NCA. In this case, the polymerization is stereoselective, ascribable to a preferable reaction between the optical enantiomorphs of the terminal residue of the growing chain and the NCA of the same chirality. On the other hand, the polymerization initiated by SarNMe₂ and MeNH(CH₂)₂CONMe₂ were stereoselective in NO₂Ph and HCONEt₂, but they were not stereoselective in m-(MeO)₂Ph. These findings indicate that the polymerizations initiated by a strong base in highly dipolar solvents are stereoselective. Apparently, the reaction between a chiral, cyclic terminal of growing chain and a chiral, cyclic activated NCA in the activated-NCA mechanism is highly stereoselective. In addition, from a kinetic investigation on on the copolymerization between L - and D -NCAs, the penultimate chiral centers were also suggested to contribute to the stereoselection. Stereoselection by the α-helical conformation of the growing chain and by a chiral, linear terminal amine have been considered so far, and the contribution from the present type of stereoselection must have been overlooked.     

Allantoin metabolism in soybean plants as influenced by grafts, a delayed inoculation with Rhizobium, and a late supply of nitrogen-compounds

Reciprocal grafts between A62-1 (nodulating variety) and A62-2(non-nodulating variety) of soybeans, delayed inoculation withRhizobium and a late supply of N-compounds to nodulated anddenodulated A62-1 plants were tested to study the regulationof allantoin production in soybeans. In the upper portions of stems of the A62-2 plants grafted ontoA62-1 plants, allantoin was accumulated in a significant quantity,but lower than the ungrafted intact A62-1 plants. The concentrationsof odier nitrogenous and sugar components were similar to thoseof the ungrafted A62-1 plants. On the other hand, in the upperstems of the A62-1 plants grafted onto A62-2 plants, littleallantoin was accumulated and the concentrations of variouscomponents were similar to those of the ungrafted intact A62-2plants. A62-1 and A62-2 plants not inoculated with Rhizobium showedapproximately the same levels of allantoin and of other componentswhen the same concentration of Ncompounds was supplied. Witha late inoculation, A62-1 plants showed a delayed accumulationof allantoin in accordance with the delayed development of nodules. When nodulated soygean plants were denodulated, the allantoinconcentration in the stems and roots rapidly decreased. Additionof ammonia, urea, or nitrate to the denodulated plants retardedthe decrease of allantoin concentration in the stems, but maintainedthe soluble Kjeldahl-N and amino-N concentrations at the samelevels as those in nodulated plants. In contrast, addition ofany one to nodulated plants did not increase the allantoin accumulation. (Received April 17, 1978; )     

Androgen metabolism in sheep

³H-Testosterone (³H-T) plus ¹⁴C-androst-4-ene-3.17-dione (A-dione) and ³H-epi-testosterone (17α-hydroxy-4-androsten-3-one) (epiT) plus ¹⁴C-T were injected intravenously into two male sheep with bile fistulae, respectively. Urine and bile samples were collected at intervals for 4–8 hours and analyzed by the use of DEAE-Sephadex A-25 and Lipidex 5000 columns, TLC, and paper chromatography; the aglycones were identified by co-crystallization with authentic standards.Five fractions were obtained from urine and bile: unconjugated, glucosiduronates, sulfates, sulfo-glucosiduronates and disulfates. In urine, the major conjugates were glucosiduronates, while sulfates predominated in bile. About 80–90% of recovered radioactivity was found to be either glucosiduronates or sulfates. Among the metabolites identified, epi-T was the principal one, accounting for 10–15% of the administered doses. Conversion to 17α-hydroxysteroids thus appears to be a major route of metabolism of the androgens administered in sheep. Other metabolites in the glucosiduronate and sulfate fractions were androsterone, etiocholanolone (3α-hydroxy-5β-androstan-17-one), 5β-androstane-3α, 17β-diol, two unknown diols and polar metabolites. The results indicated that androgen metabolism is somewhat unusual in sheep, as compared with other animals and the human.     

Solution conformation and hydrolytic activity of cyclo(D-leu-L-His)

It has been reported previously that a cyclic dipeptide, cyclo(D -Leu-L -His), showed a high hydrolytic activity toward a hydrophobic ester, p-nitrophenyl laurate. In order to determine the reason for the high catalytic activity, the conformation of cyclo(D -Leu-L -His) in aqueous solution was investigated by nuclear magnetic resonance and circular dichroism spectroscopy and compared with the conformation of cyclo(L -Leu-L -His), which was nearly inactive in otherwise the same conditions for the hydrolysis. It was demonstrated that the spatial arrangement of the hydrophobic isobutyl group of the D -leucyl residue and of the nucleophilic imidazolyl group of the L -histidyl residue in cyclo(D -Leu-L -His) matches very well with the long acyl chain and the active ester function of p-nitrophenyl laurate. On the other hand, in cyclo(L -Leu-L -His) the hydrophobic and the nucleophilic pendant groups are too close with each other to cooperate intramolecularly for the hydrolysis. It was concluded that the different steric structures of the diastereomers can explain the large difference of the catalytic activities.     

Multiple molecular forms of acid nitrophenyl phosphatase in sea urchin eggs and embryos

Acid nitrophenyl phosphatases from sea urchin eggs and embryos were investigated by gel filtration. Four different forms were found in Hemicentrotus pulcherrimus, and three forms in Anthocidaris crassispina and Pseudocentrotus depressus. The first and second forms (designated AcP-1 and AcP-2) had the highest activity in the range of pH 5.6–6.0. The third (designated AcP-3) had an apparent optimum pH between pH 4.3 and 4.8, and the fourth (designated AcP-4) showed the maximum activity at pH 3.0. AcP-1 was much more thermolabile than AcP-2 and AcP-3 at 56°C. NaF inhibited AcP-2, AcP-3, and AcP-4 but not AcP-1. AcP-1, AcP-2, and AcP-3 were observed in the three species, whereas AcP-4 was not detected in A. crassispina and P. depressus. AcP-1, AcP-2, and AcP-3 were separted by gel filtration. AcP-1 and AcP-2 of A. crassispina and H. pulcherrimus were studied in developing embryos. The behavior of these forms in gel filtration changed during development, from unfertilized eggs to the pluteus stage.