Lipase-catalyzed transamidation of non-activated amides in organic solvent

A novel biocatalytic reaction of transamidation of non-activated amides with amines is reported. Among 45 different lipolytic and proteolytic enzymes tested, only the lipase from Candida antarcticawas able to catalyze this reaction. The reaction proceeded with up to ca. 80% conversion in anhydrous methyl tert-butyl ether and worked with both N-substituted and unsubstituted amides. The biocatalytic transamidation is an equilibrium process and, therefore, higher conversions to the desired amide were achieved by using increased concentrations of the amine nucleophile.     

Studies on the toxin of Aspergillus fumigatus. XVIII. Photooxidation of asp-hemolysin in the presence of various dyes and its relation to the site of hemolytic activity

The site of hemolytic activity of a toxin isolated from Aspergillus fumigatus designated Asp-hemolysin was determined by photooxidation techniques. The hemolytic activity of this toxin was strongly inhibited by photooxidation with methylene blue, rose bengal, riboflavin, or eosin G as a sensitizer, whereas crystal violet, hematoxylin, naphthol yellow S, bromothymol blue, methyl orange, and cresol red had no effect. pH dependence of the inactivation with methylene blue was observed in the narrow range of pH values from 7.0 to 8.0, like that of the inactivation with rose bengal or riboflavin. The histidine, cysteine, methionine, tryptophan, and tyrosine content of methylene blue-photooxidized Asp-hemolysin was significantly decreased, while other amino acids were not affected. The hemolytic activity of the toxin was lost more slowly than the histidine residue, being maintained at about 50% even at the time when the histidine residue was completely lost after 30 min. Photooxidation of Asp-hemolysin in the presence of rose bengal also caused a decrease in histidine, methionine, and threonine content. These findings suggest that residues of cysteine, methionine, threonine, tryptophan, and/or tyrosine but not histidine may play an important role through stereostructure in the manifestation of the hemolytic activity of Asp-hemolysin.     

Biochemical and phenotypical correction of an envelope mutant of Salmonella typhimurium

Salmonella typhimurium DA 361 bears an env D1 mutation with the following abnormal phenotypical and biochemical characteristics: a) it autolyses at stationary phase in nutrient broth; b) it grows in chains of short rods; c) it is a poor maltose fermenter and d) it has a diphosphatidylglycerol (DPG) content twice as high than its isogenic non-lytic pair DA 362 (env D+) and LT2, of which both are derivatives. Growth of DA 361 in the presence of 400 mM ethanol leads on a 50% decrease of DPG level, thereby equalling its PG/DPG ratio with those of the control strain. Consequently, a correction on the other phenotypical and biochemical anomalies are induced since the DA 361 strain decreases its autolytic activity, ferments normally maltose and appear as rods undifferentiated from DA 362.     

Characteristics of the maintenance of the upright posture in subjects touching an external object while standing on a moving or immobile platform

Postural sway was compared for humans touching an external object while standing on an immobile or slowly moving posturographic platform. When the platform moves, the central nervous system may interpret the movement of the point of the contact with the external object as the movement of the body relative to the support or as the movement of the support itself. Thus, the information concerning the body position that is provided by the touch becomes ambiguous. It was demonstrated that contact with an external object during standing on an unstable support leads to a decrease in support sway. When a subject stands on a moving platform, this decrease is smaller than in the case of an immobile platform. Contact with an external object causes a decrease in postural responses to shank muscle vibrations on an immobile platform. On a moving platform, this decrease is nonsignificant. The change in postural sway depending on the unambiguity of afferent information is discussed in terms of the interaction between afferent signals of different modalities on the basis of the body scheme in subjects maintaining balance.Translated from Fiziologiya Cheloveka, Vol. 31, No. 1, 2005, pp. 59–65.Original Russian Text Copyright © 2005 by Kazennikov, Shlykov, Levik.     

Human skin chymotrypsin-like proteinase chymase. Subcellular localization to mast cell granules and interaction with heparin and other glycosaminoglycans

The subcellular localization of human skin chymase to mast cell granules was established by immunoelectron microscopy, and binding of chymase to the area of the dermo-epidermal junction, a basement membrane, was demonstrated immunocytochemically in cryosections incubated with purified proteinase prior to immunolabeling. Because heparin and heparan sulfate proteoglycans are major constituents of mast cell granules and basement membranes, respectively, the ability of chymase to bind to glycosaminoglycans (GAG) was investigated. Among a variety of GAGs, only binding of chymase to heparin and heparan sulfate appears physiologically significant. Binding was ionic strength-dependent, involved amino groups on the proteinase, and correlated with increasing GAG sulfate content, indicating a predominantly electrostatic association. Interaction with heparin was observed in solutions containing up to 0.5 M NaCl, and interaction with heparan sulfate was observed in solutions containing up to 0.3 M NaCl. Binding of heparin did not detectably affect catalysis of peptide substrates, but may reduce accessibility of proteinase to protein substrates. Measurements among a series of serine class proteinases indicated that heparin binding was a more common property of mast cell proteinases than proteinases stored in other secretory granules. Binding of chymase to heparin is likely to have a storage as well as a structural role within the mast cell granule, whereas binding of chymase to heparan sulfate may have physiological significance after degranulation.     

Parathyroid hormone stimulates adenylate cyclase in rat cerebral microvessels

We have studied the effect of parathyroid hormone (PTH) on adenylate cyclase of microvessels isolated from rat cerebral cortex. Native bovine (b) PTH-(1–84), the synthetic amino-terminal fragment bPTH-(1–34) and the synthetic analog [Nle⁸, Nle¹⁸, Tyr³⁴]-bPTH- (1–34) amide stimulated adenylate cyclase in a dose-dependent manner with apparent ED₅₀ values of 16 nM, 6.3 nM and 15 nM respectively. The stimulation by bPTH was greatly enhanced by guanosine triphosphate. The PTH antagonist, [Nle⁸, Nle¹⁸, Tyr³⁴]-bPTH-(3–34) amide inhibited the action of bPTH-(1–84) and bPTH-(1–34). In summary, PTH stimulated adenylate cyclase in rat cerebral microvessels in a very similar manner to its stimulation in the renal cortex.     

An apparatus to measure in vivo biomechanical behavior of dorsi- and plantarflexors of mouse ankle.

We developed an apparatus to quantify the biomechanical behavior of the dorsi- and plantarflexor muscles of the ankle of an anesthetized mouse. When the dorsi- or plantarflexor muscle group is activated by electrical stimulation of either the peroneal or tibial nerve, the apparatus measures the moment developed about the ankle during isometric, isovelocity shortening, or isovelocity lengthening contractions. Displacements may be performed over the full 105 degrees range of ankle motion with an angular resolution of 0.09 degrees. Bidirectional isovelocity ramps in ankle angle up to 1,100 degrees/s are possible and are equivalent to maximum velocities of 2.3 fiber lengths/s (Lf/s) for the fibers in tibialis anterior muscle and 11.9 Lf/s for those in gastrocnemius muscle. During single contractions of the dorsi- and plantarflexor muscle groups at 37 degrees C and with both knee and ankle joint at 90 degrees neutral position, the isometric tetanic force developed was 1.4 and 3.3 N, power output at 2.2 Lf/s was 3.1 and 5.9 mW, and power absorption at 0.5 Lf/s was 4.9 and 9.0 mW, respectively. These values are in reasonable agreement with data from the same muscle groups tested in situ. We conclude that the apparatus provides valid measurements of force and power of the dorsi- and plantarflexor muscle groups.